Functional plasticity in the substrate binding site of beta-secretase

The aspartic protease beta-secretase (BACE) cleaves the amyloid precursor protein into a 42 residue beta-peptide, which is the principal biochemical marker of Alzheimer's disease. Multiple explicit-water molecular dynamics simulations of the apo and inhibitor bound structures of BACE indicate that both open- and closed-flap conformations are accessible at room temperature and should be taken into account for inhibitor design. Correlated motion is observed within each of the two lobes of BACE, as well as for the interfacial region. A self-inhibited conformation with the side chain of Tyr71 occupying the S(1) pocket is present in some of the unbound simulations. The reversible loss of the side chain hydrogen bond between the catalytic Asp32 and Ser35, due to the concomitant reorientation of the Ser35 hydroxyl group and a water molecule conserved in pepsin-like enzymes, provides further evidence for the suggestion that Ser35 assists in proton acceptance and release by Asp32 during catalysis.     

Structural diversity of domain superfamilies in the CATH database

The CATH database of domain structures has been used to explore the structural variation of homologous domains in 294 well populated domain structure superfamilies, each containing at least three sequence diverse relatives. Our analyses confirm some previously detected trends relating sequence divergence to structural variation but for a much larger dataset and in some superfamilies the new data reveal exceptional structural variation. Use of a new algorithm (2DSEC) to analyse variability in secondary structure compositions across a superfamily sheds new light on how structures evolve. 2DSEC detects inserted secondary structures that embellish the core of conserved secondary structures found throughout the superfamily. Analysis showed that for 56% of highly populated superfamilies (>9 sequence diverse relatives), there are twofold or more increases in the numbers of secondary structures in some relatives. In some families fivefold increases occur, sometimes modifying the fold of the domain. Manual inspection of secondary structure insertions or embellishments in 48 particularly variable superfamilies revealed that although these insertions were usually discontiguous in the sequence they were often co-located in 3D resulting in a larger structural motif that often modified the geometry of the active site or the surface conformation promoting diverse domain partnerships and protein interactions. These observations, supported by automatic analysis of all well populated CATH families, suggest that accretion of small secondary structure insertions may provide a simple mechanism for evolving new functions in diverse relatives. Some layered domain architectures (e.g. mainly-beta and alpha-beta sandwiches) that recur highly in the genomes more frequently exploit these types of embellishments to modify function. In these architectures, aggregation occurs most often at the edges, top or bottom of the beta-sheets. Information on structural variability across domain superfamilies has been made available through the CATH Dictionary of Homologous Structures (DHS).     

含磷酸结合口袋的BRCT结构域功能位点预测

BRCT( BRCA1 C-terminus)是真核生物DNA损伤修复系统重要的信号传导和蛋白靶向结构域.为了探讨含磷酸结合口袋的BRCT与磷酸化配体结合的机制,对XRCC1 BRCT1、PTIP BRCT4、ECT2 BRCT1和TopBP1BRCT1进行了结构保守性和表面静电势分析.结果显示,4个BRCT的磷酸结合口袋周围所存在的结构保守并带正电势的沟槽很可能是其功能位点,并且类似的沟槽在含磷酸结合口袋的BRCT中普遍存在.沟槽两侧及底部均带有极性氨基酸残基,两侧带正电荷,而底部疏水.这说明沟槽与配体的结合以静电和疏水相互作用为主.沟槽主要位于单个BRCT中,而且4个BRCT的沟槽在形状和电荷分布上都不同,说确明BRCT配体特异性主要由单个BRCT决定.磷酸结合口袋位于沟槽中心,说明沟槽可能同时结合磷酸化残基的N端和C端附近残基.     

Functional plasticity of CH domains

With the refinement of algorithms for the identification of distinct motifs from sequence databases, especially those using secondary structure predictions, new protein modules have been determined in recent years. Calponin homology (CH) domains were identified in a variety of proteins ranging from actin cross-linking to signaling and have been proposed to function either as autonomous actin binding motifs or serve a regulatory function. Despite the overall structural conservation of the unique CH domain fold, the individual modules display a quite striking functional variability. Analysis of the actopaxin/parvin protein family suggests the existence of novel (type 4 and type 5) CH domain families which require special attention, as they appear to be a good example for how CH domains may function as scaffolds for other functional motifs of different properties.     

Structural plasticity in the oestrogen receptor ligand-binding domain

The steroid hormone receptors are characterized by binding to relatively rigid, inflexible endogenous steroid ligands. Other members of the nuclear receptor superfamily bind to conformationally flexible lipids and show a corresponding degree of elasticity in the ligand-binding pocket. Here, we report the X-ray crystal structure of the oestrogen receptor alpha (ERalpha) bound to an oestradiol derivative with a prosthetic group, ortho- trifluoromethlyphenylvinyl, which binds in a novel extended pocket in the ligand-binding domain. Unlike ER antagonists with bulky side groups, this derivative is enclosed in the ligand-binding pocket, and acts as a potent agonist. This work shows that steroid hormone receptors can interact with a wider array of pharmacophores than previously thought through structural plasticity in the ligand-binding pocket.     

Evolution of enzyme superfamilies

Enzyme evolution is often constrained by aspects of catalysis. Sets of homologous proteins that catalyze different overall reactions but share an aspect of catalysis, such as a common partial reaction, are called mechanistically diverse superfamilies. The common mechanistic steps and structural characteristics of several of these superfamilies, including the enolase, Nudix, amidohydrolase, and haloacid dehalogenase superfamilies have been characterized. In addition, studies of mechanistically diverse superfamilies are helping to elucidate mechanisms of functional diversification, such as catalytic promiscuity. Understanding how enzyme superfamilies evolve is vital for accurate genome annotation, predicting protein functions, and protein engineering.     

Functional genomics of neural and behavioral plasticity

How does the environment, particularly the social environment, influence brain and behavior and what are the underlying physiologic, molecular, and genetic mechanisms? Adaptations of brain and behavior to changes in the social or physical environment are common in the animal world, either as short-term (i.e., modulatory) or as long-term modifications (e.g., via gene expression changes) in behavioral or physiologic properties. The study of the mechanisms and constraints underlying these dynamic changes requires model systems that offer plastic phenotypes as well as a sufficient level of quantifiable behavioral complexity while being accessible at the physiological and molecular level. In this article, I explore how the new field of functional genomics can contribute to an understanding of the complex relationship between genome and environment that results in highly plastic phenotypes. This approach will lead to the discovery of genes under environmental control and provide the basis for the study of the interrelationship between an individual's gene expression profile and its social phenotype in a given environmental context.     

Functional plasticity of the BNIP-2 and Cdc42GAP Homology (BCH) domain in cell signaling and cell dynamics

The BNIP-2 and Cdc42GAP Homology (BCH) domains constitute a new and expanding family of highly conserved scaffold protein domains that regulate Rho, Ras and MAPK signaling, leading to cell growth, apoptosis, morphogenesis, migration and differentiation. Such versatility is achieved via their ability to target small GTPases and their immediate regulators such as GTPase-activating proteins (GAPs) and guanine nucleotide exchange factors (GEFs), their ability to form intra-molecular or inter-molecular interaction with itself or with other BCH domains, and also by their ability to bind diverse cellular proteins such as membrane receptors, isomerase, caspases and metabolic enzymes such as glutaminase. The presence of BCH and BCH-like domains in various proteins and their divergence from the ancestral lipid-binding CRAL-TRIO domain warrant the need to examine closely their structural, functional and regulatory plasticity in isolation or in concert with other protein modules present in the same proteins.     

Functional diversity on synaptic plasticity mediated by endocannabinoids

Endocannabinoids (eCBs) act as modulators of synaptic transmission through activation of a number of receptors, including, but not limited to, cannabinoid receptor 1 (CB1). eCBs share CB1 receptors as a common target with Δ⁹-tetrahydrocannabinol (THC), the main psychoactive ingredient in marijuana. Although THC has been used for recreational and medicinal purposes for thousands of years, little was known about its effects at the cellular level or on neuronal circuits. Identification of CB1 receptors and the subsequent development of its specific ligands has therefore enhanced our ability to study and bring together a substantial amount of knowledge regarding how marijuana and eCBs modify interneuronal communication. To date, the eCB system, composed of cannabinoid receptors, ligands and the relevant enzymes, is recognized as the best-described retrograde signalling system in the brain. Its impact on synaptic transmission is widespread and more diverse than initially thought. The aim of this review is to succinctly present the most common forms of eCB-mediated modulation of synaptic transmission, while also illustrating the multiplicity of effects resulting from specializations of this signalling system at the circuital level.     

Structural motif of phosphate-binding site common to various protein superfamilies: all-against-all structural comparison of protein-mononucleotide complexes

In order to search for a common structural motif in the phosphate-binding sites of protein-mononucleotide complexes, we investigated the structural variety of phosphate-binding schemes by an all-against-all comparison of 491 binding sites found in the Protein Data Bank. We found four frequently occurring structural motifs composed of protein atoms interacting with phosphate groups, each of which appears in different protein superfamilies with different folds. The most frequently occurring motif, which we call the structural P-loop, is shared by 13 superfamilies and is characterized by a four-residue fragment, GXXX, interacting with a phosphate group through the backbone atoms. Various sequence motifs, including Walker's A motif or the P-loop, turn out to be a structural P-loop found in a few specific superfamilies. The other three motifs are found in pairs of superfamilies: protein kinase and glutathione synthetase ATPase domain like, actin-like ATPase domain and nucleotidyltransferase, and FMN-linked oxidoreductase and PRTase.     

SSToSS--sequence-structural templates of single-member superfamilies

The presence of sequence homologues and the availability of structural information of proteins enable better understanding of the biological function of a protein family. A majority of entries in protein structural databank are single member superfamilies for which it is hard to derive motifs due to the paucity of structural homologues. Important conserved segments for these superfamilies have been identified and compiled into a database, SSToSS (Sequence Structural Templates of Single member Superfamily). Conserved regions, recognized by permitted amino acid exchanges, are mapped on the structure and various structural features (solvent accessibility, secondary structure content, hydrogen bonding and residue packing) are examined. These conserved segments with high structural feature content are projected as sequence-structural templates for the particular superfamily member. Interactive three-dimensional displays of the templates in three-dimensional structure (in Chime and RASMOL) are provided for better understanding and visualization. In SSToSS database, we also provide the application of sequence-structural templates in three different areas: multiple-motif based sequence search, multiple sequence alignment and homology modeling. In each case, the inclusion of the sequence-structural templates can give rise to sensitive and accurate results. This enables the inclusion of singletons to provide added value to the recognition of additional members, comparative modeling and in designing experiments.     

Functional plasticity of spinal circuits in long-term sports activity adaptation

Physical activity-dependent mechanisms of functional changes in spinal cord circuitry were studied in athletes involved in sports for a long time. New data were obtained for functional plasticity the cervical and lumbosacral motor spinal systems develop as a result of various long-term sports activities. Compared with nonathletes, ski racers and basketball players were found to have an extended representation area of upper- and lower-limb muscle α-motoneurons with a high reflex excitability. The area was extended by involving upper segments in all cases. Several indices of functional plasticity of the cervical and lumbar regions of the spinal cord in ski racers, who are engaged in moderate cyclic activity, were higher than in basketball players, whose movements are more various.     

Physical origins of protein superfamilies

In this work, we discovered a fundamental connection between selection for protein stability and emergence of preferred structures of proteins. Using a standard exact three-dimensional lattice model we evolve sequences starting from random ones and determine the exact native structure after each mutation. Acceptance of mutations is biased to select for stable proteins. We found that certain structures, "wonderfolds", are independently discovered numerous times as native states of stable proteins in many unrelated runs of selection. The strong dependence of lattice fold usage on the structural determinant of designability quantitatively reproduces uneven fold usage in natural proteins. Diversity of sequences that fold into wonderfold structures gives rise to superfamilies, i.e. sets of dissimilar sequences that fold into the same or very similar structures. The present work establishes a model of pre-biotic structure selection, which identifies dominant structural patterns emerging upon optimization of proteins for survival in a hot environment. Convergently discovered pre-biotic initial superfamilies with wonderfold structures could have served as a seed for subsequent biological evolution involving gene duplications and divergence.     

Active site plasticity of endonuclease V from Salmonella typhimurium

Base deamination is a major type of DNA damage under nitrosative stress. Endonuclease V initiates repair of deaminated base damage by making a nucleolytic incision one nucleotide away from the 3' side of the lesion. Within the endonuclease V family, the substrate specificities are different from one enzyme to another. In this study, we investigated deamination lesion cleavage activities of endonuclease V from the macrophage-residing pathogen, Salmonella typhimurium. Salmonella endonuclease V exhibits limited turnover on cleavage of deoxyinosine- and xanthosine-containing DNA. Binding analysis indicates that this single-turnover property is caused by tight binding to nicked products. The nicking activity is similar between the double-stranded deoxyinosine- and deoxyxanthosine-containing DNA. Cleavage rates are not affected by bases opposite the deoxyinosine or deoxyxanthosine lesions. The enzyme is also active on single-stranded deoxyinosine- and deoxyxanthosine-containing DNA. Unlike endonuclease V from Thermotoga maritima, Salmonella endonucleae V can only turnover deoxyuridine-containing DNA to a limited extent when substrate is in excess. Binding analysis indicates that Salmonella endonuclease V achieves tight binding to deoxyuridine-containing DNA, a property that distinguishes it from Thermotoga endonuclease V. Cleavage analysis on mismatch-containing DNA also indicates that the active site of Salmonella endonuclease V can accommodate pyrimidine-containing mismatches, resulting in more comparable cleavage of pyrimidine- and purine-containing mismatches. This comprehensive DNA cleavage and binding analysis reveals the plastic nature in the active site of Salmonella endonuclease V, which allows the enzyme to enfold both purine and pyrimidine deaminated lesions or base pair mismatches.     

Protein architecture: new superfamilies

Protein families whose members have evolved beyond any statistically significant similarities in sequence are sometimes called superfamilies. Structural and functional characteristics are conserved within such families and can be revealed by an analysis of protein architecture. We describe ten superfamilies that have resulted from recently determined structures; four of these are described here for the first time.     

Functional characterization of the MENTAL domain

Human metastatic lymph node (MLN) 64 is composed of two conserved regions. The amino terminus contains a conserved membrane-spanning MENTAL (MLN64 NH(2)-terminal) domain shared with an unique protein called MENTHO (MLN64 NH(2)-terminal domain homologue) and targets the protein to late endosome. The carboxyl-terminal domain is composed of a cholesterol binding steroidogenic acute regulatory-related lipid transfer domain exposed to the cytoplasm. MENTHO overexpression leads to the accumulation of enlarged endosomes. In this study, we show that MLN64 overexpression also induces the formation of enlarged endosomes, an effect that is probably mediated by the MENTAL domain. Using an in vivo photocholesterol binding assay, we find that the MENTAL domain of MLN64 is a cholesterol binding domain. Moreover, glutathione S-transferase pull-down or co-immunoprecipitation experiments demonstrate that this domain mediates homo- and hetero-interaction of MLN64 and MENTHO. In living cells, the expression of paired yellow fluorescent and cyan fluorescent fusion proteins show MENTHO homo-interaction and its interaction with MLN64. These data indicate that within late-endosomal membranes, MLN64 and MENTHO define discrete cholesterol-containing subdomains. The MENTAL domain might serve to maintain cholesterol at the membrane of late endosomes prior to its shuttle to cytoplasmic acceptor(s).     

Functional MRI of long-term potentiation: imaging network plasticity

Neurons are able to express long-lasting and activity-dependent modulations of their synapses. This plastic property supports memory and conveys an extraordinary adaptive value, because it allows an individual to learn from, and respond to, changes in the environment. Molecular and physiological changes at the cellular level as well as network interactions are required in order to encode a pattern of synaptic activity into a long-term memory. While the cellular mechanisms linking synaptic plasticity to memory have been intensively studied, those regulating network interactions have received less attention. Combining high-resolution fMRI and in vivo electrophysiology in rats, we have previously reported a functional remodelling of long-range hippocampal networks induced by long-term potentiation (LTP) of synaptic plasticity in the perforant pathway. Here, we present new results demonstrating an increased bilateral coupling in the hippocampus specifically supported by the mossy cell commissural/associational pathway in response to LTP. This fMRI-measured increase in bilateral connectivity is accompanied by potentiation of the corresponding polysynaptically evoked commissural potential in the contralateral dentate gyrus and depression of the inactive convergent commissural pathway to the ipsilateral dentate. We review these and previous findings in the broader context of memory consolidation.