Mitochondria hold diverse and pivotal roles in fundamental processes that govern cell survival, differentiation, and death, in addition to organismal growth, maintenance, and aging. The mitochondrial protein import system is a major contributor to mitochondrial biogenesis and lies at the crossroads between mitochondrial and cellular homeostasis. Recent findings highlight the mitochondrial protein import system as a signaling hub, receiving inputs from other cellular compartments and adjusting its function accordingly. Impairment of protein import, in a (...) physiological, or disease context, elicits adaptive responses inside and outside mitochondria. In this review, we discuss recent developments, relevant to the mechanisms of mitochondrial protein import regulation, with a particular focus on quality control, proteostatic and metabolic cellular responses, triggered upon impairment of mitochondrial protein import. (shrink)

The PRDM family has recently spawned considerable interest as it has been implicated in fundamental aspects of cellular differentiation and exhibits expanding ties to human diseases. The PRDMs belong to the SET domain family of histone methyltransferases, however, enzymatic activity has been determined for only few PRDMs suggesting that they act by recruiting co‐factors or, more speculatively, confer methylation of non‐histone targets. Several PRDM family members are deregulated in human diseases, most prominently in hematological malignancies and solid cancers, where they (...) can act as both tumor suppressors or drivers of oncogenic processes. The molecular mechanisms have been delineated for only few PRDMs and little is known about functional redundancy within the family. Future studies should identify target genes of PRDM proteins and the protein complexes in which PRDM proteins reside to provide a more comprehensive understanding of the biological and biochemical functions of this important protein family. (shrink)

Genetic analysis is revealing molecular components of circadian rhythms. The gene products of the period gene in Drosophila and the frequency gene in Neurospora oscillate with a circadian rhythm. A recent paper(1) has shown that the PERIOD protein can undergo both intermolecular and intramolecular interactions in vitro. The effects of temperature and two period mutations on these molecular interactions were compared to the effects of the mutations and temperature on the in vivo period length of circadian rhythms. The results (...) suggest that the molecular interactions may compete to maintain a rhythm with a constant period over a wide temperature range. (shrink)

The protein folding problem is one of the foundational problems of biochemistry and it is still considered unsolved. It basically consists of two main questions: what are the factors determining the stability of the protein’s native structure and how does the protein acquire it starting from an unfolded state. Since its first formulation, two main explanatory approaches have dominated the field of protein folding research: a thermodynamic approach focused on energetic features and a kinetic approach focused (...) on the temporal development of protein chains and structural considerations. Although these two approaches are tightly intertwined in biochemical practice and largely agree on which are the parts and activities in which the phenomenon under study should be decomposed to, there nevertheless exist important contrasts that have had repercussions on the development of the field and still engender vigorous debate. We shall analyse the historical development of the field and crucial aspects of current scientific debates. On this basis, we argue that the main sources of disagreement centre on the causal interpretation of thermodynamic and kinetic explanations, on the explanatory relevance assigned to different features of the phenomena under study and on the status of the ontological assumptions concerning the entities under study. (shrink)

Protein‐carbohydrate interactions have been found to be important in many steps in lymphocyte recirculation and inflammatory responses. A family of carbohydrate‐binding proteins or lectins, termed selectins, has been discovered and shown to be involved directly in these processes. The three known selectins, termed L‐, E‐ and P‐selectins, have domains homologous to other Ca2+‐dependent (C‐type) lectins. L‐selectin is expressed constitutively on lymphocytes, E‐selectin is expressed by activated endothelial cells, and P‐selectin is expressed by activated platelets and endothelial cells. Here, we (...) review the nature of the carbohydrate determinants in tissues recognized by these selectins. The expression of specific sialylated, fucosylated and sulfated carbohydrates in activated endothelium and high endothelial venules promotes interactions with L‐selectin on leukocyte surfaces. In contrast, E‐ and P‐selectins recognize specific carbohydrate determinants related to sialyl Lex antigen on neutrophil and monocyte surfaces. The discovery of the selectins has generated excitemient among glycoconjugate researchers that other carbohydrate‐binding proteins and their cognate ligands will be found to function in regulating many types of cellular interactions. (shrink)

The discovery and engineering of novel fluorescent proteins (FPs) from diverse organisms is yielding fluorophores with exceptional characteristics for live‐cell imaging. In particular, the development of FPs for fluorescence (or Förster) resonance energy transfer (FRET) microscopy is providing important tools for monitoring dynamic protein interactions inside living cells. The increased interest in FRET microscopy has driven the development of many different methods to measure FRET. However, the interpretation of FRET measurements is complicated by several factors including the high fluorescence (...) background, the potential for photoconversion artifacts and the relatively low dynamic range afforded by this technique. Here, we describe the advantages and disadvantages of four methods commonly used in FRET microscopy. We then discuss the selection of FPs for the different FRET methods, identifying the most useful FP candidates for FRET microscopy. The recent success in expanding the FP color palette offers the opportunity to explore new FRET pairs. (shrink)

Peroxisomal matrix proteins are synthesized on cytosolic ribosomes and rapidly transported into the organelle by a complex machinery. The data gathered in recent years suggest that this machinery operates through a syringe-like mechanism, in which the shuttling receptor PEX5 − the “plunger” − pushes a newly synthesized protein all the way through a peroxisomal transmembrane protein complex − the “barrel” − into the matrix of the organelle. Notably, insertion of cargo-loaded receptor into the “barrel” is an ATP-independent process, (...) whereas extraction of the receptor back into the cytosol requires its monoubiquitination and the action of ATP-dependent mechanoenzymes. Here, we review the main data behind this model. The peroxin PEX5 is the peroxisomal matrix protein receptor. It shuttles between the cytosol and the organelle membrane, where it gets inserted into the docking/translocation module, thus pushing the cargo-protein into the peroxisome lumen. Resetting the system involves PEX5 monoubiquitination and its extraction from the membrane by the AAA-ATPases, PEX1/PEX6. (shrink)

Membership in a protein domain database does not a domain make; a feature we realized when generating a consensus view of protein fold space with our consensus domain dictionary (CDD). This dictionary was used to select representative structures for characterization of the protein dynameome: the Dynameomics initiative. Through this endeavor we rejected a surprising 40% of the 1,695 folds in the CDD as being non‐autonomous folding units. Although some of this was due to the challenges of grouping (...) similar fold topologies, the dissonance between the cataloguing and structural qualification of protein domains remains surprising. Another potential factor is previously overlooked intrinsic disorder; predictions suggest that 40% of proteins have either local or global disorder. One thing is clear, filtering a structural database and ensuring a consistent definition for protein domains is crucial, and caution is prescribed when generalizations of globular domains are drawn from unfiltered protein domain datasets. (shrink)

Membership in a protein domain database does not a domain make; a feature we realized when generating a consensus view of protein fold space with our consensus domain dictionary (CDD). This dictionary was used to select representative structures for characterization of the protein dynameome: the Dynameomics initiative. Through this endeavor we rejected a surprising 40% of the 1,695 folds in the CDD as being non‐autonomous folding units. Although some of this was due to the challenges of grouping (...) similar fold topologies, the dissonance between the cataloguing and structural qualification of protein domains remains surprising. Another potential factor is previously overlooked intrinsic disorder; predictions suggest that 40% of proteins have either local or global disorder. One thing is clear, filtering a structural database and ensuring a consistent definition for protein domains is crucial, and caution is prescribed when generalizations of globular domains are drawn from unfiltered protein domain datasets. (shrink)

The Hippo pathway, a cascade of protein kinases that inhibits the oncogenic transcriptional coactivators YAP and TAZ, was discovered in Drosophila as a major determinant of organ size in development. Known modes of regulation involve surface proteins that mediate cell‐cell contact or determine epithelial cell polarity which, in a tissue‐specific manner, use intracellular complexes containing FERM domain and actin‐binding proteins to modulate the kinase activities or directly sequester YAP. Unexpectedly, recent work demonstrates that GPCRs, especially those signaling through Galpha12/13 (...) such as the protease activated receptor PAR1, cause potent YAP dephosphorylation and activation. This response requires active RhoA GTPase and increased assembly of filamentous (F‐)actin. Morever, cell architectures that promote F‐actin assembly per se also activate YAP by kinase‐dependent and independent mechanisms. These findings unveil the ability of GPCRs to activate the YAP oncogene through a newly recognized signaling function of the actin cytoskeleton, likely to be especially important for normal and cancerous stem cells. (shrink)

Replication protein A (RPA) is the main eukaryotic single‐stranded DNA (ssDNA) binding protein, having essential roles in all DNA metabolic reactions involving ssDNA. RPA binds ssDNA with high affinity, thereby preventing the formation of secondary structures and protecting ssDNA from the action of nucleases, and directly interacts with other DNA processing proteins. Here, we discuss recent results supporting the idea that one function of RPA is to prevent annealing between short repeats that can lead to chromosome rearrangements by (...) microhomology‐mediated end joining or the formation of hairpin structures that are substrates for structure‐selective nucleases. We suggest that replication fork catastrophe caused by depletion of RPA could result from cleavage of secondary structures by nucleases, and that failure to cleave hairpin structures formed at DNA ends could lead to gene amplification. These studies highlight the important role RPA plays in maintaining genome integrity. (shrink)

Although the importance of weak protein‐protein interactions has been understood since the 1980s, scant attention has been paid to this “quinary structure”. The transient nature of quinary structure facilitates dynamic sub‐cellular organization through loose grouping of proteins with multiple binding partners. Despite our growing appreciation of the quinary structure paradigm in cell biology, we do not yet understand how the many forces inside the cell – the excluded volume effect, the “stickiness” of the cytoplasm, and hydrodynamic interactions – (...) perturb the weakest functional protein interactions. We discuss the unresolved problem of how the forces in the cell modulate quinary structure, and to what extent the cell has evolved to exert control over the weakest biomolecular interactions. We conclude by highlighting the new experimental and computational tools coming on‐line for in vivo studies, which are a critical next step if we are to understand quinary structure in its native environment. (shrink)

Protein translocation across biological membranes is of fundamental importance for the biogenesis of organelles and in protein secretion. We will give an overview of the recent achievements in the understanding of protein translocation across mitochondrial membranes(1‐5). In particular we will focus on recently identified components of the mitochondrial import apparatus.

Although interactions of proteins with glycosaminoglycans (GAGs), such as heparin and heparan sulphate, are of great biological importance, structural requirements for protein‐GAG binding have not been well‐characterised. Ionic interactions are important in promoting protein‐GAG binding. Polyelectrolyte theory suggests that much of the free energy of binding comes from entropically favourable release of cations from GAG chains. Despite their identical charges, arginine residues bind more tightly to GAGs than lysine residues. The spacing of these residues may determine protein‐GAG (...) affinity and specificity. Consensus sequences such as XBBBXXBX, XBBXBX and a critical 20 Å spacing of basic residues are found in some protein sites that bind GAG. A new consensus sequence TXXBXXTBXXXTBB is described, where turns bring basic interacting amino acid residues into proximity. Clearly, protein‐GAG interactions play a prominent role in cell‐cell interaction and cell growth. Pathogens including virus particles might target GAG‐binding sites in envelope proteins leading to infection. BioEssays 20:156–167, 1998. © 1998 John Wiley & Sons, Inc. (shrink)

In this paper I use a case study—the discovery of the chaperon function exerted by proteins in the various steps of the hereditary process—to re-discuss the question whether the nucleic acids are the sole repositories of relevant information as assumed in the information theory of heredity. The evidence I here present of a crucial role for molecular chaperones in the folding of nascent proteins, as well as in DNA duplication, RNA folding and gene control, suggests that the family of proteins (...) acting as molecular chaperones provides information that is complementary to that stored in the nucleic acids, and equally important. A re-evaluation of the role of proteins in the hereditary process is in order away from the gene-centric approach of the information theory of heredity, to which neo-Darwinian evolutionists adhere. (shrink)

The examination of the complex cell biology of the human malaria parasite Plasmodium falciparum usually relies on the time‐consuming generation of transgenic parasites. Here, metabolic labeling and click chemistry are employed as a fast transfection‐independent method for the microscopic examination of protein S‐palmitoylation, an important post‐translational modification during the asexual intraerythrocytic replication of P. falciparum. Applying various microscopy approaches such as confocal, single‐molecule switching, and electron microscopy, differences in the extent of labeling within the different asexual developmental stages of (...) P. falciparum and the host erythrocytes over time are observed. (shrink)

Since their discovery over two decades ago, the molecular and cellular functions of the NIPSNAP family of proteins (NIPSNAPs) have remained elusive until recently. NIPSNAPs interact with a variety of mitochondrial and cytoplasmic proteins. They have been implicated in multiple cellular processes and associated with different physiologic and pathologic conditions, including pain transmission, Parkinson's disease, and cancer. Recent evidence demonstrated a direct role for NIPSNAP1 and NIPSNAP2 proteins in regulation of mitophagy, a process that is critical for cellular health and (...) maintenance. Importantly, NIPSNAPs contain a 110 amino acid domain that is evolutionary conserved from mammals to bacteria. However, the molecular function of the conserved NIPSNAP domain and its potential role in mitophagy have not been explored. It stands to reason that the highly conserved NIPSNAP domain interacts with a substrate that is ubiquitously present across all species and can perhaps act as a sensor for mitochondrial health. (shrink)

In spite of the numerous efforts made to control their transmission, parasite schistosomes still represent a serious public health concern and a major economic problem in many developing countries. Praziquantel (PZQ) is the drug of choice for the treatment of schistosomiasis and the only one that is available for mass chemotherapy. However, its widespread use and its inefficacy on juvenile parasites raise fears that schistosomes will develop drug resistance, and make the development of alternative drugs highly desirable. Protein tyrosine (...) kinases (PTKs) are key molecules that control cell differentiation and proliferation and they already represent important targets for molecular cancer therapy. The recent characterization in Schistosoma mansoni of several cytosolic and receptor PTKs, with properties similar but also divergent from their vertebrate counterparts, opens new perspectives for the development of novel strategies in chemotherapy of schistosomiasis, which could be based on the use of parasite‐specific tyrosine phosphorylation inhibitors. BioEssays 29:1281–1288, 2007. © 2007 Wiley Periodicals, Inc. (shrink)

Fatty acids (FAs) are well known to serve as substrates for reactions that provide cells with membranes and energy. In contrast to these metabolic reactions, the physiological importance of FAs themselves known as free FAs (FFAs) in cells remains obscure. Since accumulation of FFAs in cells is toxic, cells must develop mechanisms to detoxify FFAs. One such mechanism is to sequester free polyunsaturated FAs (PUFAs) into a droplet‐like structure assembled by Fas‐Associated Factor 1 (FAF1), a cytosolic protein. This sequestration (...) limits access of PUFAs to Fe2+, thereby preventing Fe2+‐catalyzed PUFA peroxidation. Consequently, assembly of the FAF1‐FFA complex is critical to protect cells from ferroptosis, a cell death pathway triggered by PUFA peroxidation. The observations that free PUFAs in cytosol are not randomly diffused but rather sequestered into a membraneless complex should open new directions to explore signaling pathways by which FFAs regulate cellular physiology. (shrink)

Coronins constitute an evolutionarily conserved family of WD‐repeat actin‐binding proteins, which can be clearly classified into two distinct groups based on their structural features. All coronins possess a conserved basic N‐terminal motif and three to ten WD repeats clustered in one or two core domains. Dictyostelium and mammalian coronins are important regulators of the actin cytoskeleton, while the fly Dpod1 and the yeast coronin proteins crosslink both actin and microtubules. Apart from that, several coronins have been shown to be involved (...) in vesicular transport. C. elegans POD‐1 and Drosophila coro regulate the actin cytoskeleton, but also govern vesicular trafficking as indicated by mutant phenotypes. In both organisms, defects in cytoskeleton and trafficking lead to severe developmental defects ranging from abnormal cell division to aberrant formation of morphogen gradients. Finally, mammalian coronin 7 appears not to execute any cytoskeleton‐related functions, but rather participates in regulating Golgi trafficking. Here, we review recent data providing more insight into molecular mechanisms underlying the regulation of F‐actin structures, cytoskeletal rearrangements and intracellular membrane transport by coronin proteins and the way that they might link cytoskeleton with trafficking in development and disease. BioEssays 27:625–632, 2005. © 2005 Wiley Periodicals, Inc. (shrink)

The lateral mobility of membrane lipids and proteins is presumed to play an important functional role in biomembranes. Photobleaching studies have shown that many proteins in the plasma membrane have diffusion coefficients at least an order of magnitude lower than those obtained when the same proteins are reconstituted in artificial bilayer membranes. Depending on the protein, it has been shown that either the cytoplasmic domain or the ectodomain is the key determinant of its lateral mobility. Single particle tracking microscopy, (...) which allows the motions of single or small groups of membrane molecules to be followed, promises not only to reveal new features of membrane dynamics, but also to help explain longstanding puzzles presented by the photobleaching studies, particularly the so‐called immobile fraction. The combination of the two complementary technologies should measurably enhance our understanding of membrane microstructure. (shrink)

This paper considers two recent arguments that structure should not be regarded as the fundamental individuating property of proteins. By clarifying both what it might mean for certain properties to play a fundamental role in a classification scheme and the extent to which structure plays such a role in protein classification, I argue that both arguments are unsound. Because of its robustness, its importance in laboratory practice, and its explanatory centrality, primary structure should be regarded as the fundamental distinguishing (...) characteristic of protein taxonomy. (shrink)

RNA binding proteins (RBPs) are key factors for the regulation of gene expression by binding to cis elements, i.e. short sequence motifs in RNAs. Recent studies demonstrate that cooperative binding of multiple RBPs is important for the sequence‐specific recognition of RNA and thereby enables the regulation of diverse biological activities by a limited set of RBPs. Cross‐linking immuno‐precipitation (CLIP) and other recently developed high‐throughput methods provide comprehensive, genome‐wide maps of protein‐RNA interactions in the cell. Structural biology gives detailed insights (...) into molecular mechanisms and principles of RNA recognition by RBPs, but has so far focused on single RNA binding proteins and often on single RNA binding domains. The combination of high‐throughput methods and detailed structural biology studies is expected to greatly advance our understanding of the code for protein‐RNA recognition in gene regulation, as we review in this article. (shrink)

A protein kinase cascade is involved in the action of some mitogens. The cascade begins with receptor tyrosine kinase activation by growth factors. The resulting signal is transmitted into cells via phospholipid metabolism which produces a variety of second messengers and by intracellular protein kinase activation. The signal is then propagated and disseminated via a network of other proteln kinases and protein phosphatases. Recent research suggests that ribosomal protein S6 kinase and casein kinase II are two (...) important elements in the kinase cascade that leads to the initiation of growth. The nature and some properties of these hitherto lesser known enzymes is considered. (shrink)

In this article two questions are discussed with regard to semiosis in protein biosynthesis for nano engines. (1) What kind of semiosis is involved in the construction of these proteins? and (2) How can we explain the semiotic process observed? With regard to the first issue we draw attention to comparisons between semiosis in protein biosynthesis and human natural language. The notion of normativity appears to be of great importance for both. A comparison also demonstrates differences. Nevertheless, because (...) of the normative symbolic information processing in it, we suggest to employ the term symbolic reference (employed in linguistics as a distinguishing feature of human language) to indicate the semiotic processes in protein biosynthesis. With regard to explaining semiosis in protein synthesis we compare different approaches. We conclude that a Kantian approach should be preferred. In such an approach strengths of the mechanistic and organicist approaches can be combined, and the observed symbolic information processing acknowledged. (shrink)

The model organism Saccharomyces cerevisiae is providing new insights into the molecular and cellular changes that are related to aging. The yeast protein Sir2p (Silent Information Regulator 2) is a histone deacetylase involved in transcriptional silencing and the control of genomic stability. Recent results have led to the identification of Sir2p as a crucial determinant of yeast life span. Dosage, intracellular localization, and activity of Sir2p all have important effects on yeast longevity. For instance, calorie restriction apparently increases yeast (...) life span by increasing Sir2p activity. Since Sir2p‐related proteins have been identified in many prokaryotic and eukaryotic organisms, the fundamental principles derived from the studies in yeast may prove valuable in directing our future research toward an understanding of the mechanisms of aging in higher eukaryotes. BioEssays 23:327–332, 2001. © 2001 John Wiley & Sons, Inc. (shrink)

To explore whether the generation of new protein folds could be linked to metallic cofactor recruitment, we identified the oldest examples of folds for manganese, iron, zinc, and copper proteins by analyzing their fold‐domain mapping patterns. We discovered that the generation of these folds was tightly coupled to corresponding metals. We found that the emerging order for these folds, i.e., manganese and iron protein folds appeared earlier than zinc and copper counterparts, coincides with the putative bioavailability of the (...) corresponding metals in the ancient anoxic ocean. Therefore, we conclude that metallic cofactors, like organic cofactors, play an evolutionary role in the formation of new protein folds. This link could be explained by the emergence of protein structures with novel folds that could fulfill the new protein functions introduced by the metallic cofactors. These findings not only have important implications for understanding the evolutionary mechanisms of protein architectures, but also provide a further interpretation for the evolutionary story of superoxide dismutases. (shrink)

Lipid droplets (LDs) are ubiquitous, neutral lipid storage organelles that act as hubs of metabolic processes. LDs are structurally unique with a hydrophobic core that mainly consists of neutral lipids, sterol esters, and triglycerides, enclosed within a phospholipid monolayer. Nascent LD formation begins with the accumulation of neutral lipids in the endoplasmic reticulum (ER) bilayer. The ER membrane proteins such as seipin, LDAF1, FIT, and MCTPs are reported to play an important role in the formation of nascent LDs. As the (...) LDs grow, they unmix from the highly charged ER membrane to form mature LDs. LD biogenesis is an exciting, emerging research area, and herein, we discuss the recent progress in our understanding of the formation of eukaryotic nascent LDs. We focus on the role of ER membrane shaping proteins such as reticulons and reticulon‐like proteins, membrane lipids, and cytoskeleton proteins such as septin in the formation of nascent LDs. (shrink)

In metazoans, the extracellular matrix (ECM) provides a dynamic, heterogeneous microenvironment that has important supportive and instructive roles. Although the primary site of action of ECM proteins is extracellular, evidence is emerging for non‐canonical intracellular roles. Examples include osteopontin, thrombospondins, IGF‐binding protein 3 and biglycan, and relate to roles in transcription, cell‐stress responses, autophagy and cancer. These findings pose conceptual problems on how proteins signalled for secretion can be routed to the cytosol or nucleus, or can function in environments (...) with diverse redox, pH and ionic conditions. We review evidence for intracellular locations and functions of ECM proteins, and current knowledge of the mechanisms by which they may enter intracellular compartments. We evaluate the experimental methods that are appropriate to obtain rigorous evidence for intracellular localisation and function. Better insight into this under‐researched topic is needed to decipher the complete spectrum of physiological and pathological roles of ECM proteins. -/- . (shrink)

Rho/Rac proteins constitute a subgroup of the Ras superfamily of GTP hydrolases. Although originally implicated in the control of cytoskeletal events, it is currently known that these GTPases coordinate diverse cellular functions, including cell polarity, vesicular trafficking, the cell cycle and transcriptomal dynamics. In this review, we will provide an overview on the recent advances in this field regarding the mechanism of regulation and signaling, and the roles in vivo of this important GTPase family. BioEssays 29:356–370, 2007. © 2007 Wiley (...) Periodicals, Inc. (shrink)

In the 1950s and 1960s, an interfield interaction between molecular biologists and biochemists integrated important discoveries about the mechanism of protein synthesis. This extended discovery episode reveals two general reasoning strategies for eliminating gaps in descriptions of the productive continuity of mechanisms: schema instantiation and forward chaining/backtracking. Schema instantiation involves filling roles in an overall framework for the mechanism. Forward chaining and backtracking eliminate gaps using knowledge about types of entities and their activities. Attention to mechanisms highlights salient features (...) of this historical episode while providing general reasoning strategies for mechanism discovery. (shrink)

Around 1960, leading figures in the international conservation circuit – such as Julian Huxley, Frank Fraser Darling and E. Barton Worthington – successfully propagated new visions about the value of undomesticated African mammals. Against traditional ideas, they presented these mammals as a highly efficient source of protein for growing African populations. In line with this vision, they challenged non-interventionist ideals of nature preservation, and launched proposals for active management through game ‘ranching’ and ‘cropping’. As such, they created a new (...) socio-technical imaginary for Africa's future, in which the consumption of wildlife meat took up a central position. This article explores the motivations of Western conservationists for this drastic rebranding. It argues that the rationale of considering African wildlife in terms of protein played an important symbolical role at various levels. It was crucial in the reorganization of the transnational networks of conservation, but also in the boosting of their scientific reputation, in the restructuring of their institutional ties, and in their attempts to maintain an authoritative position for Western ecologists in a rapidly decolonizing world. (shrink)

The prediction of protein–protein interactions based on independently obtained structural information for each interacting partner remains an important challenge in computational chemistry. Procedures where hypothetical interaction models (or decoys) are generated, then ranked using a biochemically relevant scoring function have been garnering interest as an avenue for addressing such challenges. The program PatchDock has been shown to produce reasonable decoys for modeling the association between pig alpha-amylase and the VH-domains of camelide antibody raised against it. We designed a (...) biochemically relevant method by which PatchDock decoys could be ranked in order to separate near-native structures from false positives. Several thousand steps of energy minimization were used to simulate induced fit within the otherwise rigid decoys and to rank them. We applied a partial free energy function to rank each of the binding modes, improving discrimination between near-native structures and false positives. Sorting decoys according to strain energy increased the proportion of near-native decoys near the bottom of the ranked list. Additionally, we propose a novel method which utilizes regression analysis for the selection of minimization convergence criteria and provides approximation of the partial free energy function as the number of algorithmic steps approaches infinity. (shrink)

BET proteins such as Brd3 and Brd4 are chromatin‐associated factors, which control gene expression programs that promote inflammation and cancer. The Nrf2 transcription factor is a master regulator of genes that protect the organism against xenobiotic attack and oxidative stress. Nrf2 has demonstrated anti‐inflammatory activity and can support cancer cell malignancy. This review describes the discovery, mechanism and biomedical implications of the regulatory interplay between Nrf2 and BET proteins. Both Nrf2 and BET proteins are established drug targets. Small molecules that (...) either activate or suppress these proteins are currently tested in clinical trials. The crosstalk between Nrf2 and BET proteins may have important, and until now overlooked, implications for the therapeutic effects of these drugs. Based on the information covered in this review, it should be possible to design combinatorial treatment strategies for cancer and inflammatory diseases, which may improve the efficacy of targeting a Nrf2 or BET proteins individually. (shrink)

Lysosomal positioning is an important factor in regulating cellular responses, including autophagy. Because proteins encoded by disease‐responsible genes are involved in lysosomal trafficking, proper intracellular lysosomal trafficking is thought to be essential for cellular homeostasis. In the past few years, the mechanisms of lysosomal trafficking have been elucidated with a focus on adapter proteins linking motor proteins to lysosomes. Here, we outline recent findings on the mechanisms of lysosomal trafficking by focusing on adapter protein c‐Jun NH2‐terminal kinase‐interacting protein (...) (JIP) 4, which plays a central role in this process, and other JIP4 functions and JIP family proteins. Additionally, we discuss neuronal diseases associated with aberrance in the JIP family protein. Accumulating evidence suggests that chemical manipulation of lysosomal positioning may be a therapeutic approach for these neuronal diseases. (shrink)

Protein complexes perform many important functions in the cell. Large‐scale studies of protein–protein interactions have not only revealed new complexes but have also placed many proteins into multiple complexes. Whilst the advocates of hypothesis‐free research touted the discovery of these shared components as new links between diverse cellular processes, critical commentators denounced many of the findings as artefacts, thus questioning the usefulness of large‐scale approaches. Here, we survey proteins known to be shared between complexes, as established in (...) the literature, and compare them to shared components found in high‐throughput screens. We discuss the various challenges to the identification and functional interpretation of bona fide shared components, namely contaminants, variant and megacomplexes, and transient interactions, and suggest that many of the novel shared components found in high‐throughput screens are neither the results of contamination nor central components, but appear to be primarily regulatory links in cellular processes. BioEssays 26:1333–1343, 2004. © 2004 Wiley Periodicals, Inc. (shrink)

Protein misfolding is a topic that is of primary interest both in biology and medicine because of its impact on fundamental processes and disease. In this review, we revisit the concept of protein misfolding and discuss how the field has evolved from the study of globular folded proteins to focusing mainly on intrinsically unstructured and often disordered regions. We argue that this shift of paradigm reflects the more recent realisation that misfolding may not only be an adverse event, (...) as originally considered, but also may fulfil a basic biological need to compartmentalise the cell with transient reversible granules. We nevertheless provide examples in which structure is an important component of a much more complex aggregation behaviour that involves both structured and unstructured regions of a protein. We thus suggest that a more comprehensive evaluation of the mechanisms that lead to aggregation might be necessary. (shrink)

Here I discuss findings that suggest a universal mechanism for proteins (and RNA) to recognize and interact with various binding partners by selectively binding to different conformations that pre‐exist in the free protein's conformational ensemble. The tandem RNA recognition motif domains of splicing factor U2AF65 fluctuate in solution between a predominately closed conformation in which the RNA binding site of one of the domains is blocked, and a lowly populated open conformation in which both RNA binding pockets are accessible. (...) RNA binding to U2AF65 may thus occur through the weakly populated open conformation, and the binding interaction stabilizes the open conformation. The conformational diversity observed in U2AF65 might also facilitate binding to diverse RNA sequences as found in the polypyrimidine tracts that help define 3′ splice sites. Similar binding pathways in other systems have important consequences in biological regulation, molecular evolution, and information storage. (shrink)

The rel family of proteins can be defined as a group of proteins that share sequence homology over a 300 amino acid region termed the rel domain. The rel family comprises important regulatory proteins from a wide variety of species and includes the Drosophila morphogen dorsal, the mammalian transcription factor NF‐kB, the avian oncogene v‐rel, and the cellular proto‐oncogene c‐rel. Over the last two years it has become apparent that these proteins function as DNA‐binding transcription factors, and that their activity (...) is regulated at the level of subcellular localization. (shrink)

The prediction of the secondary structure of a protein from its amino acid sequence is an important step towards the prediction of its three-dimensional structure. However, the accuracy of ab initio secondary structure prediction from sequence is about 80 % currently, which is still far from satisfactory. In this study, we proposed a novel method that uses binomial distribution to optimize tetrapeptide structural words and increment of diversity with quadratic discriminant to perform prediction for protein three-state secondary structure. (...) A benchmark dataset including 2,640 proteins with sequence identity of less than 25 % was used to train and test the proposed method. The results indicate that overall accuracy of 87.8 % was achieved in secondary structure prediction by using ten-fold cross-validation. Moreover, the accuracy of predicted secondary structures ranges from 84 to 89 % at the level of residue. These results suggest that the feature selection technique can detect the optimized tetrapeptide structural words which affect the accuracy of predicted secondary structures. (shrink)

The best‐characterized model pathway of protein folding, that of disulphide bond formation in the small protein BPTI, has been questioned recently. A reinvestigation of that pathway, using alternative methods, concluded that the intermediates with non‐native disulphide bonds accumulated to lower levels than previously had been observed(17). On this basis, a revised pathway was proposed that simply omitted those intermediates. Even if totally correct, however, the new observations are not inconsistent with the important characteristics of the original pathway and (...) even confirmed many of them. Certain crucial observations that were the experimental basis for the original pathway were ignored, and these observations invalidate the revised pathway. (shrink)

AbstractmRNA stability is increasingly recognized as being essential for controlling the expression of a wide variety of transcripts during neuronal development and synaptic plasticity. In this context, the role of AU‐rich elements (ARE) contained within the 3′ untranslated region (UTR) of transcripts has now emerged as key because of their high incidence in a large number of cellular mRNAs. This important regulatory element is known to significantly modulate the longevity of mRNAs by interacting with available stabilizing or destabilizing RNA‐binding proteins (...) (RBP). Thus, in parallel with the emergence of ARE, RBP are also gaining recognition for their pivotal role in regulating expression of a variety of mRNAs. In the nervous system, the member of the Hu family of ARE‐binding proteins known as HuD, has recently been implicated in multiple aspects of neuronal function including the commitment and differentiation of neuronal precursors as well as synaptic remodeling in mature neurons. Through its ability to interact with ARE and stabilize multiple transcripts, HuD has now emerged as an important regulator of mRNA expression in neurons. The present review is designed to provide a comprehensive and updated view of HuD as an RBP in the nervous system. Additionally, we highlight the role of HuD in multiple aspects of a neuron's life from early differentiation to changes in mature neurons during learning paradigms and in response to injury and regeneration. Finally, we describe the current state of knowledge concerning the molecular and cellular events regulating the expression and activity of HuD in neurons. BioEssays 28: 822–833, 2006. © 2006 Wiley Periodicals, Inc. (shrink)

Cytoplasmically synthesized proteins are directed into chloroplasts by amino terminal transit sequences of the precursor proteins. For proteins of the thylakoid lumen, transit sequences are also important in directing proteins to the lumen.

A cell's biochemistry is now known to be the biochemistry of molecular machines, that is, protein complexes that are assembled and dismantled in particular locations within the cell as needed. One important element in our understanding has been the ability to begin to see where proteins are in cells and what they are doing as they go about their business. Accordingly, there is now a strong impetus to discover new ways of looking at the workings of proteins in living (...) cells. Although the use of fluorescent tags to track individual proteins in cells has a long history, the availability of laser-based confocal microscopes and the imaginative exploitation of the green fluorescent protein from jellyfish have provided new tools of great diversity and utility. It is now possible to watch a protein bind its substrate or its partners in real time and with submicron resolution within a single cell. The importance of processes of self-organisation represented by protein folding on the one hand and subcellular organelles on the other are well recognised. Self-organisation at the intermediate level of multimeric protein complexes is now open to inspection. BioEssays 22:180–187, 2000. ©2000 John Wiley & Sons, Inc. (shrink)

Iron uptake and storage in mammalian cells is at least partly regulated at a posttranscriptional level by the iron regulatory proteins (IRP‐1 and IRP‐2). These cytoplasmic regulators share 79% similarity in protein sequence and bind tightly to conserved mRNA stem‐loops, named iron‐responsive elements (IREs). The IRP:IRE interaction underlies the regulation of translation and stability of several mRNAs central to iron metabolism. The question of why the cell requires two such closely related regulatory proteins may be resloved as we learn (...) more about the expression and regulation of these proteins. It is evident so far that, despite similarities, the IRPs differ in several important respects. They are coordinately regulated by cellular iron, but whereas IRP‐1 is inactivated by high iron levels, IRP‐2 is rapidly degraded. Further differences arise in their expression and RNA‐binding specificity. The two proteins each recognise a large repertoire of IRE‐like sequences, including a small group of exclusive RNA targets. These findings hint that IRP‐1 and IRP‐2 may bind preferenitially to certain mRNAs in vivo, possibly extending their known functions beyond the regulation of intracellular iron homeostasis. (shrink)

Saethre‐Chotzen syndrome (SCS), a human autosomal dominant condition with limb defects and craniosynostosis, is caused by haploinsufficiency of TWIST1, a basic helix–loop–helix (bHLH) transcription factor. Until recently, the molecular pathogenesis of the limb defects in SCS has not been well understood. Now, Firulli et al.1 show in mouse and chick that ectopic expression of a related bHLH protein, Hand2, results in phenocopies of the limb defects caused by Twist1 loss‐of‐function mutations. These two proteins interact in a dosage‐dependent antagonistic manner, (...) and both can be regulated through phosphorylation at conserved helix I amino acid residues. These findings provide an important link between the misregulation of Twist1 dimerization and the limb phenotypes observed in SCS. BioEssays 27:1102–1106, 2005. © 2005 Wiley Periodicals, Inc. (shrink)

A crucially important part of the biosphere – the virosphere – is too often overlooked. Inclusion of the virosphere into the global picture of protein structure space reveals that 63 protein domain superfamilies in viruses do not have any structural and evolutionary relatives in modern cellular organisms. More than half of these have functions which are not virus‐specific and thus might be a source of new folds and functions for cellular life. The number of viruses on the planet (...) exceeds that of cells by an order of magnitude and viruses evolve up to six orders of magnitude faster. As a result, cellular species are subject to a constitutive ‘flow‐through’ of new viral genetic material. Due to this and the relaxed evolutionary constraints in viruses, the transfer of domains between host‐to‐virus could be a mechanism for accelerated protein evolution. The virosphere could be an engine for the genesis of protein structures, and may even have been so before the last universal common ancestor of cellular life. (shrink)

The correct execution of the DNA replication process is crucially import for the maintenance of genome integrity of the cell. Several types of sources, both endogenous and exogenous, can give rise to DNA damage leading to the DNA replication fork arrest. The processes by which replication blockage is sensed by checkpoint sensors and how the pathway leading to resolution of stalled forks is activated are still not completely understood. However, recent emerging evidence suggests that one candidate for a sensor (...) of replication stress is ATR and that, together with a member of RecQ family helicases, Werner syndrome protein (WRN) and MRE11 complex, can collaborate to promote the restarting of DNA synthesis through the resolution of stalled replication forks. Here, we discuss how WRN, the MRE11 complex and the ATR kinase could work together in response to replication blockage to avoid DNA replication fork collapse and genome instability. BioEssays 26:306–313, 2004. © 2004 Wiley Periodicals, Inc. (shrink)

The order of discovery can have a profound effect upon the way in which we think about the function of a gene. In E. coli, recA is nearly essential for cell survival in the presence of DNA damage. However, recA was originally identified, as a gene required to obtain recombinant DNA molecules in conjugating bacteria. As a result, it has been frequently assumed that recA promotes the survival of bacteria containing DNA damage by recombination in which DNA strand exchanges occur. (...) We now know that several of the processes that interact with or are controlled by recA, such as excision repair and translesion synthesis, operate to ensure that DNA replication occurs processively without strand exchanges. Yet the view persists in the literature that recA functions primarily to promote recombination during DNA repair. With the benefit of hindsight and more than three decades of additional research, we reexamine some of the classical experiments that established the concept of DNA repair by recombination, and we consider the possibilities that recombination is not an efficient mechanism for rescuing damaged cells, and that recA may be important for maintaining processive replication in a manner that does not generally promote recombination. BioEssays 23:463–470, 2001. © 2001 John Wiley & Sons, Inc. (shrink)

We now know the structures of over 200 proteins to atomic resolution. Despite the impressive extent and quality of the results, crystal‐structure analysis has often been thought of as limited in scope, not only in its restriction to samples that can be crystallized, but in the more important respect that taking ‘snapshots’ of proteins does not directly address the complex spatio‐temporal organization of the processes in which proteins participate. It is suggested here that, as the field has matured, this second (...) limitation is gradually being overcome. As we gain increased access to structures of proteins in different conformational states – for example, in conformations produced by different states of ligation – and to families of homologous proteins, we can proceed from the statics of protein structure to the dynamics of conformational change, function, and evolution.A new scientific speciality has grown up around the solved structures: it has as its goal the elucidation of general principles of protein structure and function, to provide a theoretical framework for understanding the properties of proteins revealed by experiment. In this article we shall discuss some of the activity in this field. It will emerge clearly, I believe, that the increasing number and variety of solved structures is exerting a cumulative force. General principles are emerging from comparisons of related proteins and contrasts of dissimilar ones: the whole corpus of data is greater than the sum of the parts. (shrink)