Molecular Recognition Feature Research Articles

MEETING REPORT: THE SEVENTH CONFERENCE OF THE HELLENIC SOCIETY FOR COMPUTATIONAL BIOLOGY AND BIOINFORMATICS

The Hellenic Society For Computational Biology and Bioinformatics (HSCBB) was conceived as a trans-national society, representing two countries that share the same language and history, Cyprus and Greece. The first, unofficial meeting was held in Heraklion in 2006, followed by Athens in 2007 and Thessalonica in 2008. After the formation of the Society and its ‘inaugural’, fourth scientific conference in Athens during 2009, the HSCBB organized two meetings in Alexandroupolis and Patras, with the aspiration to provide the necessary exposure of graduate students to new developments, expand its horizons towards the international community, promote the development of the field and formulate a solid agenda for the future, all consistent with its primary goals (http://www.hscbb.gr). The HSCBB12 meeting, which took place in Heraklion confirmed that, even in a period of austerity and societal crisis, the local community is increasingly active. Thanks to the support of national and international sponsors, including the Computational and Structural Biotechnology Journal, the HSCBB12 has been an unprecedented success. Approximately 100 participants have registered, providing for the first time a much clearer picture of activities of the bioinformatics communities in the two countries and material for a mapping exercise undertaken by the HSCBB, as a vehicle for further growth. One verdict from this analysis has been that today, unlike seven years ago, all major academic centers in the two countries have acquired some bioinformatics activity, with a wide range of expertise, much depending on the local needs of the corresponding organizations. Four keynote speakers provided a wider perspective of translational bioinformatics in their own special fields: Manolis Dermitzakis (University of Geneva) on human population genomics, Anton Enright (European Bioinformatics Institute) on miRNA computational genomics, Alice McHardy (Max-Planck-Institut fur Informatik) on human influenza A virus evolution, and Alexis Stamatakis (Heidelberg Institute for Theoretical studies) on computational challenges in biodiversity informatics. Moreover, another four invited speakers covered diverse, yet more practical areas in the field: Graham Ball (Nottingham Trent University) on pathway modelling, Guy Cochrane (European Bioinformatics Institute) on the European Nucleotide Archive, Ioannis Kontodinas (Biomax Informatics) on knowledge management for systems biology, and Kitsos Louis (IMBB-FORTH) on the role of ontologies in modern biology. All these contributions were highly appreciated by their audiences. The topics covered by the conference included the following areas: structural bioinformatics (interaction site analysis, sequence-based structural classification, molecular recognition feature analysis); sequence analysis (transcriptome analysis, SNP analysis, structural variomics); systems biology (stochastic modeling of protein dynamics, metabolomics, epigenomics); gene regulation and miRNA (miRNA profiling, miRNA duplex prediction, miRNA ontologies); algorithms, methods & tools (tree pruning algorithms, GPU accelerators, scientific workflow management systems); applied/translational bioinformatics (cell signaling simulations, text mining, evolutionary bioinformatics, data integration, biomarker discovery); genome and comparative biology (gene duplication detection, microbial genome annotation, pangenome analysis). The list of the 70 or so contributions (20 oral presentations, 38 posters, 4 invited presentations, 1 three-day workshop, plus the 8 keynote and invited lectures) has exceeded expectations for another reason. For the first time, contributions at the HSCBB conference had a significant number of co-authorships with multiple, highly prestigious institutions, including the EBI, the Joint Genome Institute at Berkeley, and Imperial College London. In total, according to one study published a few years ago in the HSCBB10 proceedings, 400 papers from 63 institutions have been published in one generation, with an exponential increase during the past decade, indicating that bioinformatics have now taken root in Greece and Cyprus, with a new generation of scientists acquiring the expertise and the ability to instigate new research and education programs. According to another bibliometric study, also published in the HSCBB10 proceedings, there is still scope for growth both in quantity and quality. Greece can potentially increase the number of bioinformatics laboratories from a dozen or so today to over 30 while Cyprus might also develop bioinformatics from a handful of teams to twice that number. Moreover, the impact of HSCBB in the literature is comparable to that of Australia and New Zealand, if the GDP per capita rank of the two pairs of countries is taken into account. In other words, Greece and Cyprus have reached a level where their computational biology communities arguably have a respectable standing on the world stage. As pointed out elsewhere, the field has now grown to a point that biological research is virtually impossible without computation, so that even smaller countries do have a role in this collective effort worldwide (http://www.ploscollections.org/article/browseIssue.action?issue=info%3Adoi%2F10.1371%2Fissue.pcol.v03.i03). We hope that the effort of the HSCBB members will ultimately generate the interest of the international community and establish a presence beyond the borders of Greece and Cyprus.

Identification of a molecular recognition feature in the E1A oncoprotein that binds the SUMO conjugase UBC9 and likely interferes with polySUMOylation

Hub proteins have central roles in regulating cellular processes. By targeting a single cellular hub, a viral oncogene may gain control over an entire module in the cellular interaction network that is potentially comprised of hundreds of proteins. The adenovirus E1A oncoprotein is a viral hub that interacts with many cellular hub proteins by short linear motifs/molecular recognition features (MoRFs). These interactions transform the architecture of the cellular protein interaction network and virtually reprogram the cell. To identify additional MoRFs within E1A, we screened portions of E1A for their ability to activate yeast pseudohyphal growth or differentiation. This identified a novel functional region within E1A conserved region 2 comprised of the sequence EVIDLT. This MoRF is necessary and sufficient to bind the N-terminal region of the SUMO conjugase UBC9, which also interacts with SUMO noncovalently and is involved in polySUMOylation. Our results suggest that E1A interferes with polySUMOylation, but not with monoSUMOylation. These data provide the first insight into the consequences of the interaction of E1A with UBC9, which was initially described in 1996. We further demonstrate that polySUMOylation regulates pseudohyphal growth and promyelocytic leukemia body reorganization by E1A. In conclusion, the interaction of the E1A oncogene with UBC9 mimics the normal binding between SUMO and UBC9 and represents a novel mechanism to modulate polySUMOylation.

N-terminal Domains of DELLA Proteins Are Intrinsically Unstructured in the Absence of Interaction with GID1/Gibberellic Acid Receptors

The plant growth-repressing DELLA proteins (DELLAs) are known to represent a convergence point in integration of multiple developmental and environmental signals in planta, one of which is hormone gibberellic acid (GA). Binding of the liganded GA receptor (GID1/GA) to the N-terminal domain of DELLAs is required for GA-induced degradation of DELLAs via the ubiquitin-proteasome pathway, thus derepressing plant growth. However, the conformational changes of DELLAs upon binding to GID1/GA, which are the key to understanding the precise mechanism of GID1/GA-mediated degradation of DELLAs, remain unclear. Using biophysical, biochemical, and bioinformatics approaches, we demonstrated for the first time that the unbound N-terminal domains of DELLAs are intrinsically unstructured proteins under physiological conditions. Within the intrinsically disordered N-terminal domain of DELLAs, we have identified several molecular recognition features, sequences known to undergo disorder-to-order transitions upon binding to interacting proteins in intrinsically unstructured proteins. In accordance with the molecular recognition feature analyses, we have observed the binding-induced folding of N-terminal domains of DELLAs upon interaction with AtGID1/GA. Our results also indicate that DELLA proteins can be divided into two subgroups in terms of their molecular compactness and their interactions with monoclonal antibodies.

Unfoldomics of Human Genetic Diseases: Illustrative Examples of Ordered and Intrinsically Disordered Members of the Human Diseasome

Intrinsically disordered proteins (IDPs) constitute a recently recognized realm of atypical biologically active proteins that lack stable structure under physiological conditions, but are commonly involved in such crucial cellular processes as regulation, recognition, signaling and control. IDPs are very common among proteins associated with various diseases. Recently, we performed a systematic bioinformatics analysis of the human diseasome, a network that linked the human disease phenome (which includes all the human genetic diseases) with the human disease genome (which contains all the disease-related genes) (Goh, K. I., Cusick, M. E., Valle, D., Childs, B., Vidal, M., and Barabasi, A. L. (2007). The human disease network. Proc. Natl. Acad. Sci. U.S.A. 104, 8685-90). The analysis of this diseasome revealed that IDPs are abundant in proteins linked to human genetic diseases, and that different genetic disease classes varied dramatically in the IDP content (Midic U., Oldfield C.J., Dunker A.K., Obradovic Z., Uversky V.N. (2009) Protein disorder in the human diseasome: Unfoldomics of human genetic diseases. BMC Genomics. In press). Furthermore, many of the genetic disease-related proteins were shown to contain at least one molecular recognition feature, which is a relatively short loosely structured protein region within a mostly disordered segment with the feature gaining structure upon binding to a partner. Finally, alternative splicing was shown to be abundant among the diseasome genes. Based on these observations the human-genetic-disease-associated unfoldome was created. This minireview describes several illustrative examples of ordered and intrinsically disordered members of the human diseasome.

A RasGTP-Induced Conformational Change in C-RAF Is Essential for Accurate Molecular Recognition

The dysregulation of Ras-RAF signaling is associated with many types of human cancer. However, the kinetic and dynamic features of the mutual molecular recognition of Ras and RAF remain unknown. Here, we developed a technique for imaging single-pair fluorescence resonance energy transfer in living cells, and coupled this technique to single-molecule kinetic analysis to investigate how C-RAF (a subtype of RAF) molecules distinguish the active form of Ras (RasGTP) from the inactive form (RasGDP). Functional fragments of C-RAF containing the Ras-binding domains did not detect the switch in Ras activity in living cells as efficiently as did C-RAF. Single-molecule analysis showed that RasGDP associates with closed-conformation C-RAF, whereas the association of C-RAF with RasGTP immediately triggers the open RAF conformation, which induces an effective interaction between C-RAF and RasGTP. Spontaneous conformational changes from closed C-RAF to the open form rarely occur in quiescent cells. The conformational change in C-RAF is so important to Ras-RAF molecular recognition that C-RAF mutants lacking the conformational change cannot distinguish between RasGDP and RasGTP. The manipulation of the conformation of an effector molecule is a newly identified function of RasGTP.

Introduction to the Molecular Recognition and Self-Assembly Special Feature

Molecular recognition is a mature branch of chemical science, and why shouldn't it be? Decades of studies in physical organic chemistry have defined and evaluated the weak intermolecular forces involved when 2 molecules encounter each other. Every bimolecular reaction, whether it occurs in the gas phase, dilute solution, or an enzyme's interior, begins with a recognition event. It is an apt time to explore recognition in a larger context, that of multicomponent assemblies, and this special issue of 14 contributions is a beginning.

Structural and functional studies on Ribonuclease S, retro S and retro-inverso S peptides

Ribonuclease S peptide and S protein offer a unique complementation system to understand the finer features of molecular recognition. In the present study the S peptide (1–16), and its retro and retro-inverso analogs have been analyzed for their structural and biological attributes. RPHPLC, CD, and NMR analyses have revealed that the physicochemical and conformational properties of the S peptide are distinct from those of its retro and retro-inverso analogs. On the functional side, while the S peptide complemented the S protein to give RNase activity, was recognized by anti-S peptide antibodies and induced T cell proliferation, neither the retro nor the retro-inverso S peptides could do so.

Abundance of Intrinsic Disorder in Protein Associated with Cardiovascular Disease

Evidence that many protein regions and even entire proteins lacking stable tertiary and/or secondary structure in solution (i.e., intrinsically disordered proteins) might be involved in protein-protein interactions, regulation, recognition, and signal transduction is rapidly accumulating. These signaling proteins play a crucial role in the development of several pathological conditions, including cancer. To test a hypothesis that intrinsic disorder is also abundant in cardiovascular disease (CVD), a data set of 487 CVD-related proteins was extracted from SWISS-PROT. CVD-related proteins are depleted in major order-promoting residues (Trp, Phe, Tyr, Ile, and Val) and enriched in some disorder-promoting residues (Arg, Gln, Ser, Pro, and Glu). The application of a neural network predictor of natural disordered regions (PONDR VL-XT) together with cumulative distribution function (CDF) analysis, charge-hydropathy plot (CH plot) analysis, and alpha-helical molecular recognition feature (alpha-MoRF) indicator revealed that CVD-related proteins are enriched in intrinsic disorder. In fact, the percentage of proteins with 30 or more consecutive residues predicted by PONDR VL-XT to be disordered was 57 +/- 4% for CVD-associated proteins. This value is close that described earlier for signaling proteins (66 +/- 6%) and is significantly larger than the content of intrinsic disorder in eukaryotic proteins from SWISS-PROT (47 +/- 4%) and in nonhomologous protein segments with a well-defined three-dimensional structure (13 +/- 4%). Furthermore, CDF and CH-plot analyses revealed that 120 and 36 CVD-related proteins, respectively, are wholly disordered. This high level of intrinsic disorder could be important for the function of CVD-related proteins and for the control and regulation of processes associated with cardiovascular disease. In agreement with this hypothesis, 198 alpha-MoRFs were predicted in 101 proteins from the CVD data set. A comparison of disorder predictions with the experimental structural and functional data for a subset of the CVD-associated proteins indicated good agreement between predictions and observations. Thus, our data suggest that intrinsically disordered proteins might play key roles in cardiovascular disease.

Intrinsic Disorder in Transcription Factors

Intrinsic disorder (ID) is highly abundant in eukaryotes, which reflect the greater need for disorder-associated signaling and transcriptional regulation in nucleated cells. Although several well-characterized examples of intrinsically disordered proteins in transcriptional regulation have been reported, no systematic analysis has been reported so far. To test for the general prevalence of intrinsic disorder in transcriptional regulation, we used the predictor of natural disorder regions (PONDR) to analyze the abundance of intrinsic disorder in three transcription factor datasets and two control sets. This analysis revealed that from 94.13 to 82.63% of transcription factors possess extended regions of intrinsic disorder, relative to 54.51 and 18.64% of the proteins in two control datasets, which indicates the significant prevalence of intrinsic disorder in transcription factors. This propensity of transcription factors to intrinsic disorder was confirmed by cumulative distribution function analysis and charge-hydropathy plots. The amino acid composition analysis showed that all three transcription factor datasets were substantially depleted in order-promoting residues and significantly enriched in disorder-promoting residues. Our analysis of the distribution of disorder within the transcription factor datasets revealed that (a) the AT-hooks and basic regions of transcription factor DNA-binding domains are highly disordered; (b) the degree of disorder in transcription factor activation regions is much higher than that in DNA-binding domains; (c) the degree of disorder is significantly higher in eukaryotic transcription factors than in prokaryotic transcription factors; and (d) the level of alpha-MoRF (molecular recognition feature) prediction is much higher in transcription factors. Overall, our data reflected the fact that eukaryotes with well-developed gene transcription machinery require transcription factor flexibility to be more efficient.

Anticalins versus antibodies: made-to-order binding proteins for small molecules

Engineering proteins to bind small molecules presents a challenge as daunting as drug discovery, for both hinge upon our understanding of receptor–ligand molecular recognition. However, powerful techniques from combinatorial molecular biology can be used to rapidly select artificial receptors. While traditionally researchers have relied upon antibody technologies as a source of new binding proteins, the lipocalin scaffold has recently emerged as an adaptable receptor for small molecule binding. ‘Anticalins’, engineered lipocalin variants, offer some advantages over traditional antibody technology and illuminate features of molecular recognition between receptors and small molecule ligands.

New supramolecular transition metal catalysts

AbstractThe covalent connection of a catalytically active transition metal center with a water‐soluble receptor (host molecule) generates a new type of supramolecular catalyst in which the features of molecular recognition, phase transfer catalysis and transition metal catalysis are combined in a single system. The first examples of this principle make use of commercially available β‐cyclodextrin (β‐CD) as the receptor and rhodium complexes of diphosphanes as the catalytically active center, these being covalently connected to one another via a spacer. In competitive hydrogenation of certain olefins, unusual degrees of substrate selectivity based on molecular recognition are observed, not possible by conventional transition metal catalysts. The two‐phase (water/organic) hydrogenation of nitro‐aromatics also is a smooth process catalyzed by these supramolecular complexes. They also constitute an unusually active catalyst system for the selective hydroformylation of higher olefins such as 1‐octene in a two‐phase system. Dendrimers having diphosphane moieties on the surface provide ligands for transition metals, the corresponding metal complexes (e.g., Pd) functioning as efficient catalysts which can be recycled due to their nanoscopic properties.

Supramolecular transition metal catalysts in two-phase systems

The concept of covalently connecting a catalytically active transition metal center with a water-soluble receptor (host molecule) makes a new type of supramolecular catalysis possible in which the features of molecular recognition, phase transfer catalysis and transition metal catalysis are combined in a single system. The first examples of this principle make use of the commercially available β-cyclodextrin (β-CD) as the receptor and rhodium complexes of diphosphanes as the catalytically active center, these being covalently connected to one another via a spacer. In competitive hydrogenation of certain olefins unusual degrees of substrate selectivity based on the molecular recognition are observed, not possible by conventional transition metal catalysts. The two-phase (H 2O/organic) hydrogenation of nitro-aromatics also is a smooth process with these supramolecular catalysts. They also constitute an unusually active catalyst system for the selective hydroformylation of higher olefins such as 1-octene in a two-phase system.

Novel cyclic analogs of angiotensin II with cyclization between positions 5 and 7: conformational and biological implications.

To study the conformational features of molecular recognition of angiotensin II (Asp1-Arg2-Val3-Tyr4-Val/IIe5-His6-Pro7-Phe8, AII), the synthesis and biological testing of several cyclic analogs of AII cyclized between positions 5 and 7 have been performed. The synthesized analogs were Sar1-Arg2-Val3-Tyr4-cyclo(Cys5-His6-Pen7)-Phe8 (3), Sar1-Arg2-Val3-Tyr4-cyclo(Asp5-His6-Apt7)-Phe8 (4), Sar1-Arg2-Val3-Tyr4-cyclo(Glu5-His6-Apt7)-Phe8 (5), Sar1-Arg2-Val3-Tyr4-cyclo-(Cys5-His6-Mpt7)-Phe8 (6), Sar1-Arg2-Val3-Tyr4-cyclo(Cys5-His6-Mpc7)-Phe8 (7), Sar1-Arg2-Val3-Tyr4-cyclo(Hcy5-His6-Mpt7)-Phe8 (8), and Sar1-Arg2-Val3-Tyr4-cyclo(Hcy5-His6-Mpc7)-Phe8 (9), where Apt stands for 4-amino-trans-proline, and Mpt and Mpc for 4-mercapto-trans- and -cis-prolines, respectively. Compound (9) showed good affinity at AT-1 receptors, namely a KD = 20 nM. In functional assays, it showed the characteristics of a weak partial agonist with a relative affinity of 0.26% of that for AII and an intrinsic efficacy, alpha E, of 0.42. Molecular modeling suggested a possible explanation for this finding: the relatively strong binding and the weak partial agonistic activity of compound 9 are due to interaction with AT-1 receptor of only two functionally important groups, namely, the side chains of the His6 and Phe8 residues.

A molecular mechanics analysis of molecular recognition by cyclodextrin mimics of α‐chymotrypsin

AbstractThe method of molecular mechanics is used to investigate the structural and electrostatic features of molecular recognition by β‐cyclodextrin and capped β‐cyclodextrin models of α‐chymotrypsin. Since capped β‐cyclodextrin has been shown to be the more effective biomimetic catalyst, these features of molecular recognition can be interpreted in terms of the relationship between molecular structure and catalytic function. Calculations in vacuo show that the addition of an N‐methylformamide “cap” substituent to each glucose unit appears to change the relative orientation of some glucose fragments from that found in the X‐ray structure of the β‐cyclodextrin macrocycle. These results indicate that certain structural components of molecular recognition, such as the orientation of the secondary hydroxyls and the related orientation of the caps, may be implicated in the catalysis. In addition, the electrostatic component of molecular recognition was investigated by the analysis of molecular electrostatic potential maps calculated in planes parallel to the average plane of the glycosidic oxygen atoms. The results indicate that the addition of the caps to the β‐cyclodextrin macrocycle subtly alters the pattern of the maps in each plane. However, the general qualitative features of electrostatic recognition by β‐cyclodextrin and capped β‐cyclodextrin are similar.

Electrostatic features of molecular recognition by cyclic urea mimics of chymotrypsin

The molecular electrostatic-potential pattern was used to investigate the electrostatic features of molecular recognition by two cyclic urea mimics of the active site of α-chymotrypsin. The structures of the mimics were obtained by molecular-mechanics evaluation of the conformational potential-energy surface of the molecules. Calculations were done by using two different atomic point-charge sets in order to assess the effect of charge on the electrostatic potential pattern. The molecules studies were: (1) a “full” mimic of chymotrypsin containing the hydroxyl, imidazole, and carboxylate anion functionalities typical of the active site of the enzyme, and (2) a “partial” mimic with only the hydroxyl and imidazole functional groups. Comparison of the molecular electrostatic-potential patterns of the two mimics in both charge sets showed that the largest differences were due to the structural addition of the carboxylate anion, rather than any particular differences in the choice of atomic point charge. For the full mimic, the pattern was essentially dominated by the negative charge on the carboxylate. Small structural changes which optimized the orientation of the catalytic components had little effect on the electrostatic potential pattern of the molecule. In the absence of the anionic functionality, greater differences were noted in the electrostatic potential pattern of the partial mimic in the two charge sets. The choice of atomic point charge was seen to influence the hydrogen-bonding pattern of the hydroxyl and imidazole moieties, resulting in differences in the spatial orientation of the electrostatic potential minima. In general, both charge sets produced molecular electrostatic-potential patterns which indicated that long-range electrostatic interactions would direct the cationic end of the substrate into the electron-rich binding site. However, specific local features of the electrostatic potential pattern were found to depend on point-charge set through the influence of charge on the hydrogen-bonding pattern.