Beta propellers: structural rigidity and functional diversity

Recently solved structures and proposed models have helped to reveal the structural characteristics of the beta-propeller fold, as well as the features that contribute to its high rigidity and stability. Possible strategies for identifying beta-propeller proteins in newly characterised sequences are helping to overcome the problems of predicting the beta-propeller fold from amino acid sequences.     

Actin: from structural plasticity to functional diversity

This article addresses the multiple activities of actin. Starting out with the history of actin's discovery, purification and structure, it emphasizes the close relation between structure and function. In this context, we also point to unconventional actin conformations. Their existence in living cells is not yet well documented, however, they seem to play a special role in the supramolecular patterning that underlies some of the physiological functions of actin. Conceivably, such conformations may contribute to actin's diverse activities in the nucleus that are poorly understood so far.     

The guanidino-group modifying enzymes: structural basis for their diversity and commonality

The guanidino-group modifying enzyme (GME) superfamily contains many drug targets, including metabolic enzymes from pathogenic microorganisms as well as key regulatory proteins from higher eukaryotes. These enzymes, despite their diverse sequences, adopt the common alpha/beta propeller fold and catalyze the modification of (methylated) guanidino groups. Our structural superposition and structure-based alignment for the GMEs have identified key residues that are involved in the catalysis and substrate binding. We have shown that conserved guanidino-carboxyl interactions are utilized in two different ways; the acidic residues in the catalytic site form hydrogen bonds to the substrate guanidino group, and the enzyme Arg residues at several key positions recognize the carboxyl group of the substrate and fix its orientation. Based on this observation, we have proposed rules for classifying the GME sequences and predicting their molecular function from the conservation of the key acidic and Arg residues. Other novel motifs have been identified, which involve residues that are not in direct contact with the substrate but are likely to stabilize the active-site conformation through hydrogen-bonding networks. In addition, we have examined the domain architecture of the GMEs. Although most members consist of a single catalytic domain, fold recognition analysis has identified a likely bifunctional enzyme from a cyanobacterium. It has also revealed common immunoglobulin-like beta-sandwich domains found in the enzymes that recognize protein substrates. These findings will be useful for predicting the precise mechanism of action for potential novel targets and designing therapeutic compounds against them.     

Evolution: structural and functional diversity of cadherin at the adherens junction

Adhesion between cells is essential to the evolution of multicellularity. Indeed, morphogenesis in animals requires firm but flexible intercellular adhesions that are mediated by subcellular structures like the adherens junction (AJ). A key component of AJs is classical cadherins, a group of transmembrane proteins that maintain dynamic cell-cell associations in many animal species. An evolutionary reconstruction of cadherin structure and function provides a comprehensive framework with which to appreciate the diversity of morphogenetic mechanisms in animals.     

Adaptive evolution of multiple-variable exons and structural diversity of drug-metabolizing enzymes

Background  

The human genome contains a large number of gene clusters with multiple-variable-first exons, including the drug-metabolizing UDP glucuronosyltransferase (UGT1) and I-branching β-1,6-N-acetylglucosaminyltransferase (GCNT2, also known as IGNT) clusters, organized in a tandem array, similar to that of the protocadherin (PCDH), immunoglobulin (IG), and T-cell receptor (TCR) clusters. To gain insight into the evolutionary processes that may have shaped their diversity, we performed comprehensive comparative analyses for vertebrate multiple-variable-first-exon clusters.     

Expression of virus-encoded proteinases: functional and structural similarities with cellular enzymes.

Many viruses express their genome, or part of their genome, initially as a polyprotein precursor that undergoes proteolytic processing. Molecular genetic analyses of viral gene expression have revealed that many of these processing events are mediated by virus-encoded proteinases. Biochemical activity studies and structural analyses of these viral enzymes reveal that they have remarkable similarities to cellular proteinases. However, the viral proteinases have evolved unique features that permit them to function in a cellular environment. In this article, the current status of plant and animal virus proteinases is described along with their role in the viral replication cycle. The reactions catalyzed by viral proteinases are not simple enzyme-substrate interactions; rather, the processing steps are highly regulated, are coordinated with other viral processes, and frequently involve the participation of other factors.     

β Propellers: structural rigidity and functional diversity

Recently solved structures and proposed models have helped to reveal the structural characteristics of the β-propeller fold, as well as the features that contribute to its high rigidity and stability. Possible strategies for identifying β-propeller proteins in newly characterised sequences are helping to overcome the problems of predicting the β-propeller fold from amino acid sequences.     

The C2 domain calcium-binding motif: structural and functional diversity.

The C2 domain is a Ca(2+)-binding motif of approximately 130 residues in length originally identified in the Ca(2+)-dependent isoforms of protein kinase C. Single and multiple copies of C2 domains have been identified in a growing number of eukaryotic signalling proteins that interact with cellular membranes and mediate a broad array of critical intracellular processes, including membrane trafficking, the generation of lipid-second messengers, activation of GTPases, and the control of protein phosphorylation. As a group, C2 domains display the remarkable property of binding a variety of different ligands and substrates, including Ca2+, phospholipids, inositol polyphosphates, and intracellular proteins. Expanding this functional diversity is the fact that not all proteins containing C2 domains are regulated by Ca2+, suggesting that some C2 domains may play a purely structural role or may have lost the ability to bind Ca2+. The present review summarizes the information currently available regarding the structure and function of the C2 domain and provides a novel sequence alignment of 65 C2 domain primary structures. This alignment predicts that C2 domains form two distinct topological folds, illustrated by the recent crystal structures of C2 domains from synaptotagmin 1 and phosphoinositide-specific phospholipase C-delta 1, respectively. The alignment highlights residues that may be critical to the C2 domain fold or required for Ca2+ binding and regulation.     

Glycine receptors: recent insights into their structural organization and functional diversity

Strychnine-sensitive glycine receptors (GlyRs) are known to mediate synaptic inhibition in spinal cord, brainstem and other regions of the CNS. During the past 5 years, considerable progress has been made in delineating structural determinants of ligand binding and channel activation in recombinant GlyRs. Furthermore, immunohistochemical and gene inactivation studies have disclosed distinct distributions and functions of differentially expressed GlyR subtypes in retina, hippocampus and the dorsal horn of the spinal cord. Accordingly, GlyRs regulate not only the excitability of motor and sensory neurones, but are also essential for the processing of photoreceptor signals, neuronal development and inflammatory pain sensitization. Hence, these receptors constitute promising targets for the development of clinically useful compounds.     

Cytochromes P450: exploiting diversity and enabling application as biocatalysts

The remarkable chemical reactivity and substrate range displayed by cytochromes P450 (P450s) renders them attractive as potential catalysts for a host of challenging chemical reactions in industry. The opportunities afforded by these biocatalysts are increased by the availability of greater diversity provided by the genomic resource and the variant libraries of well-known P450s produced by rational and random engineering techniques. The exploitation of this enormous diversity will require novel tools in screening, to identify enzyme reactions of interest, and also in the enabling of these valuable activities through protein engineering and bioprocess optimisation.     

Revealing the structural and functional diversity of plant cell walls

The extensive knowledge of the chemistry of isolated cell wall polymers, and that relating to the identification and partial annotation of gene families involved in their synthesis and modification, is not yet matched by a sophisticated understanding of the occurrence of the polymers within cell walls of the diverse cell types within a growing organ. Currently, the main sets of tools that are used to determine cell-type-specific configurations of cell wall polymers and aspects of cell wall microstructures are antibodies, carbohydrate-binding modules (CBMs) and microspectroscopies. As these tools are applied we see that cell wall polymers are extensively developmentally regulated and that there is a range of structurally distinct primary and secondary cell walls within organs and across species. The challenge now is to document cell wall structures in relation to diverse cell biological events and to integrate this knowledge with the emerging understanding of polymer functions.     

Molecular and structural basis of metabolic diversity mediated by prenyldiphosphate converting enzymes

General thermodynamic calculations using the semiempiric PM3 method have led to the conclusion that prenyldiphosphate converting enzymes require at least one divalent metal cation for the activation and cleavage of the diphosphate–prenyl ester bond, or they must provide structural elements for the efficient stabilization of the intermediate prenyl cation. The most important common structural features, which guide the product specificity in both terpene synthases and aromatic prenyl transferases are aromatic amino acid side chains, which stabilize prenyl cations by cation–π interactions. In the case of aromatic prenyl transferases, a proton abstraction from the phenolic hydroxyl group of the second substrate will enhance the electron density in the phenolic ortho-position at which initial prenylation of the aromatic compound usually occurs.A model of the structure of the integral transmembrane-bound aromatic prenyl transferase UbiA was developed, which currently represents the first structural insight into this group of prenylating enzymes with a fold different from most other aromatic prenyl transferases. Based on this model, the structure–activity relationships and mechanistic aspects of related proteins, for example those of Lithospermum erythrorhizon or the enzyme AuaA from Stigmatella aurantiaca involved in the aurachin biosynthesis, were elucidated. The high similarity of this group of aromatic prenyltransferases to 5-epi-aristolochene synthase is an indication of an evolutionary relationship with terpene synthases (cyclases). This is further supported by the conserved DxxxD motif found in both protein families. In contrast, there is no such relationship to the aromatic prenyl transferases with an ABBA-fold, such as NphB, or to any other known family of prenyl converting enzymes. Therefore, it is possible that these two groups might have different evolutionary ancestors.     

Bacterial whole-cell biocatalysts by surface display of enzymes: toward industrial application

Despite the first report on the bacterial display of a recombinant peptide appeared almost 30 years ago, industrial application of cells with surface-displayed enzymes is still limited. To display an enzyme on the surface of a living cell bears several advantages. First of all, neither the substrate nor the product of the enzymatic reaction needs to cross a membrane barrier. Second, the enzyme being linked to the cell can be separated from the reaction mixture and hence the product by simple centrifugation. Transfer to a new substrate preparation results in multiple cycles of enzymatic conversion. Finally, the anchoring in a matrix, in this case, the cell envelope stabilizes the enzyme and makes it less accessible to proteolytic degradation and material adsorption resulting in continuous higher activities. These advantages in common need to balance some disadvantages before this application can be taken into account for industrial processes, e.g., the exclusion of the enzyme from the cellular metabolome and hence from redox factors or other co-factors that need to be supplied. Therefore, this digest describes the different systems in Gram-positive and Gram-negative bacteria that have been used for the surface display of enzymes so far and focuses on examples among these which are suitable for industrial purposes or for the production of valuable resources, not least in order to encourage a broader application of whole-cell biocatalysts with surface-displayed enzymes.     

Phospholipase A(2) enzymes: structural diversity in lipid messenger metabolism

The phospholipases A(2) (PLA(2)s) are a large family of enzymes with varied lipidic products which are involved in numerous signal transduction pathways. The structural and functional characterization of several PLA(2)s have revealed the various mechanisms used by these enzymes to ingeniously manipulate the phospholipidic metabolic machinery.     

Aldehyde dehydrogenases: widespread structural and functional diversity within a shared framework.

Sequences of 16 NAD and/or NADP-linked aldehyde oxidoreductases are aligned, including representative examples of all aldehyde dehydrogenase forms with wide substrate preferences as well as additional types with distinct specificities for certain metabolic aldehyde intermediates, particularly semialdehydes, yielding pairwise identities from 15 to 83%. Eleven of 23 invariant residues are glycine and three are proline, indicating evolutionary restraint against alteration of peptide chain-bending points. Additionally, another 66 positions show high conservation of residue type, mostly hydrophobic residues. Ten of these occur in predicted beta-strands, suggesting important interior-packing interactions. A single invariant cysteine residue is found, further supporting its catalytic role. A previously identified essential glutamic acid residue is conserved in all but methyl malonyl semialdehyde dehydrogenase, which may relate to formation by that enzyme of a CoA ester as a product rather than a free carboxylate species. Earlier, similarity to a GXGXXG segment expected in the NAD-binding site was noted from alignments with fewer sequences. The same region continues to be indicated, although now only the first glycine residue is strictly conserved and the second (usually threonine) is not present at all, suggesting greater variance in coenzyme-binding interactions.     

Natural and amyloid self-assembly of S100 proteins: structural basis of functional diversity

The S100 proteins are 10-12 kDa EF-hand proteins that act as central regulators in a multitude of cellular processes including cell survival, proliferation, differentiation and motility. Consequently, many S100 proteins are implicated and display marked changes in their expression levels in many types of cancer, neurodegenerative disorders, inflammatory and autoimmune diseases. The structure and function of S100 proteins are modulated by metal ions via Ca(2+) binding through EF-hand motifs and binding of Zn(2+) and Cu(2+) at additional sites, usually at the homodimer interfaces. Ca(2+) binding modulates S100 conformational opening and thus promotes and affects the interaction with p53, the receptor for advanced glycation endproducts and Toll-like receptor 4, among many others. Structural plasticity also occurs at the quaternary level, where several S100 proteins self-assemble into multiple oligomeric states, many being functionally relevant. Recently, we have found that the S100A8/A9 proteins are involved in amyloidogenic processes in corpora amylacea of prostate cancer patients, and undergo metal-mediated amyloid oligomerization and fibrillation in vitro. Here we review the unique chemical and structural properties of S100 proteins that underlie the conformational changes resulting in their oligomerization upon metal ion binding and ultimately in functional control. The possibility that S100 proteins have intrinsic amyloid-forming capacity is also addressed, as well as the hypothesis that amyloid self-assemblies may, under particular physiological conditions, affect the S100 functions within the cellular milieu.