Two decades of thyroglobulin type-1 domain research

Thyroglobulin type-1 repeats are primarily found in thyroglobulin and several other functionally unrelated proteins. Because a few of them exhibit inhibitory activity against cysteine proteases they were named thyropins (thyroglobulin type-1 domain protease inhibitors). In contrast to cystatins, the best-characterized group of papain-like protease inhibitors, they exhibit greater selectivity in their interactions with target proteases. Interestingly, a few members inhibit aspartic protease cathepsin D and metalloproteases. In contrast to the inhibitory fragment of the major histocompatibility complex class II-associated p41 form of invariant chain, whose structural integrity appears mandatory for its inhibitory properties, short polypeptides derived from insulin-like growth factor-binding proteins exhibit the same activity as the structure of the whole fragment. Taken together, the results indicate that the thyroglobulin type-1 repeat is a structural motif occasionally employed as an inhibitor of proteases.     

Dual concentration-dependent activity of thyroglobulin type-1 domain of testican: specific inhibitor and substrate of cathepsin L

The thyroglobulin type-1 (Tg-1) domain is a protein module that occurs in a variety of secreted and membrane proteins and is recognised as a potent inhibitor of cysteine peptidases. We present here some properties of the Tg-1 domain of human testican, a modularly organised proteoglycan secreted mainly by brain cells, the exact in vivo function of which is not yet clear. The domain was prepared as a recombinant protein in a Pichia pastoris expression system and its activity was demonstrated by specific and selective inhibition of cathepsin L (K(i) =0.14 nM). Interaction at high enzyme and inhibitor concentrations resulted in degradation of the domain by cathepsin L, which was not observed under conditions used for the determination of kinetic parameters. No inhibitory activity could be detected for cathepsin K, but it exhibited a very similar degradation pattern. Homology modelling provided a good explanation for the different behaviour observed with the two enzymes. Firstly, the steric fit between the interfaces of testican domain and cathepsin L is stabilised by numerous favourable forces, while no such interactions are evident in the complex with cathepsin K, and repulsive interactions even prevent access of the domain to the active site of papain. Secondly, the prolonged first loop of the domain occupies a position near the catalytic cysteine residue in a more substrate-like manner, enabling cleavage of the Gly22-Ala23 bond.     

Structure,function and evolution of the hemerythrin‐like domain superfamily

Hemerythrin‐like proteins have generally been studied for their ability to reversibly bind oxygen through their binuclear nonheme iron centers. However, in recent years, it has become increasingly evident that some members of the hemerythrin‐like superfamily also participate in many other biological processes. For instance, the binuclear nonheme iron site of YtfE, a hemerythrin‐like protein involved in the repair of iron centers in Escherichia coli, catalyzes the reduction of nitric oxide to nitrous oxide, and the human F‐box/LRR‐repeat protein 5, which contains a hemerythrin‐like domain, is involved in intracellular iron homeostasis. Furthermore, structural data on hemerythrin‐like domains from two proteins of unknown function, PF0695 from Pyrococcus furiosus and NMB1532 from Neisseria meningitidis, show that the cation‐binding sites, typical of hemerythrin, can be absent or be occupied by metal ions other than iron. To systematically investigate this functional and structural diversity of the hemerythrin‐like superfamily, we have collected hemerythrin‐like sequences from a database comprising fully sequenced proteomes and generated a cluster map based on their all‐against‐all pairwise sequence similarity. Our results show that the hemerythrin‐like superfamily comprises a large number of protein families which can be classified into three broad groups on the basis of their cation‐coordinating residues: (a) signal‐transduction and oxygen‐carrier hemerythrins (H‐HxxxE‐HxxxH‐HxxxxD); (b) hemerythrin‐like (H‐HxxxE‐H‐HxxxE); and, (c) metazoan F‐box proteins (H‐HExxE‐H‐HxxxE). Interestingly, all but two hemerythrin‐like families exhibit internal sequence and structural symmetry, suggesting that a duplication event may have led to the origin of the hemerythrin domain.     

Diversity, taxonomy and evolution of medium-chain dehydrogenase/reductase superfamily.

A comprehensive, structural and functional, in silico analysis of the medium-chain dehydrogenase/reductase (MDR) superfamily, including 583 proteins, was carried out by use of extensive database mining and the blastp program in an iterative manner to identify all known members of the superfamily. Based on phylogenetic, sequence, and functional similarities, the protein members of the MDR superfamily were classified into three different taxonomic categories: (a) subfamilies, consisting of a closed group containing a set of ideally orthologous proteins that perform the same function; (b) families, each comprising a cluster of monophyletic subfamilies that possess significant sequence identity among them and might share or not common substrates or mechanisms of reaction; and (c) macrofamilies, each comprising a cluster of monophyletic protein families with protein members from the three domains of life, which includes at least one subfamily member that displays activity related to a very ancient metabolic pathway. In this context, a superfamily is a group of homologous protein families (and/or macrofamilies) with monophyletic origin that shares at least a barely detectable sequence similarity, but showing the same 3D fold. The MDR superfamily encloses three macrofamilies, with eight families and 49 subfamilies. These subfamilies exhibit great functional diversity including noncatalytic members with different subcellular, phylogenetic, and species distributions. This results from constant enzymogenesis and proteinogenesis within each kingdom, and highlights the huge plasticity that MDR superfamily members possess. Thus, through evolution a great number of taxa-specific new functions were acquired by MDRs. The generation of new functions fulfilled by proteins, can be considered as the essence of protein evolution. The mechanisms of protein evolution inside MDR are not constrained to conserve substrate specificity and/or chemistry of catalysis. In consequence, MDR functional diversity is more complex than sequence diversity. MDR is a very ancient protein superfamily that existed in the last universal common ancestor. It had at least two (and probably three) different ancestral activities related to formaldehyde metabolism and alcoholic fermentation. Eukaryotic members of this superfamily are more related to bacterial than to archaeal members; horizontal gene transfer among the domains of life appears to be a rare event in modern organisms.     

Individual recombinant thyroglobulin type-1 domains are substrates for lysosomal cysteine proteinases

Thyroglobulin contains 11 repeats of a motif called thyroglobulin type-1 domain that show sequence similarity to some proteins exhibiting inhibitory activity against cysteine proteinases. Here we report that thyroglobulin decreases the activity of cathepsins B, H, L, and papain. To examine the possible involvement of particular type-1 domains in that decrease of activity, some individual thyroglobulin type-1 domains were expressed in E. coli. These recombinant domains proved to be substrates for cathepsins B, H, L, and papain instead of inhibitors. The cleavage points with cathepsins B and L on the second and the fourth domains were determined. The possible reasons for degradation are discussed.     

Bioinformatics of the TULIP domain superfamily

Proteins of the BPI (bactericidal/permeability-increasing protein)-like family contain either one or two tandem copies of a fold that usually provides a tubular cavity for the binding of lipids. Bioinformatic analyses show that, in addition to its known members, which include BPI, LBP [LPS (lipopolysaccharide)-binding protein)], CETP (cholesteryl ester-transfer protein), PLTP (phospholipid-transfer protein) and PLUNC (palate, lung and nasal epithelium clone) protein, this family also includes other, more divergent groups containing hypothetical proteins from fungi, nematodes and deep-branching unicellular eukaryotes. More distantly, BPI-like proteins are related to a family of arthropod proteins that includes hormone-binding proteins (Takeout-like; previously described to adopt a BPI-like fold), allergens and several groups of uncharacterized proteins. At even greater evolutionary distance, BPI-like proteins are homologous with the SMP (synaptotagmin-like, mitochondrial and lipid-binding protein) domains, which are found in proteins associated with eukaryotic membrane processes. In particular, SMP domain-containing proteins of yeast form the ERMES [ER (endoplasmic reticulum)-mitochondria encounter structure], required for efficient phospholipid exchange between these organelles. This suggests that SMP domains themselves bind lipids and mediate their exchange between heterologous membranes. The most distant group of homologues we detected consists of uncharacterized animal proteins annotated as TM (transmembrane) 24. We propose to group these families together into one superfamily that we term as the TULIP (tubular lipid-binding) domain superfamily.     

Molecular evolution of the cadherin superfamily

This review deals with the large and pleiotropic superfamily of cadherins and its molecular evolution. We compiled literature data and an in-depth phylogenetic analysis of more than 350 members of this superfamily from about 30 species, covering several but not all representative branches within metazoan evolution. We analyzed the sequence homology between either ectodomains or cytoplasmic domains, and we reviewed protein structural data and genomic architecture. Cadherins and cadherin-related molecules are defined by having an ectodomain in which at least two consecutive calcium-binding cadherin repeats are present. There are usually 5 or 6 domains, but in some cases as many as 34. Additional protein modules in the ectodomains point at adaptive evolution. Despite the occurrence of several conserved motifs in subsets of cytoplasmic domains, these domains are even more diverse than ectodomains and most likely have evolved separately from the ectodomains. By fine tuning molecular classifications, we reduced the number of solitary superfamily members. We propose a cadherin major branch, subdivided in two families and 8 subfamilies, and a cadherin-related major branch, subdivided in four families and 11 subfamilies. Accordingly, we propose a more appropriate nomenclature. Although still fragmentary, our insight into the molecular evolution of these remarkable proteins is steadily growing. Consequently, we can start to propose testable hypotheses for structure-function relationships with impact on our models of molecular evolution. An emerging concept is that the ever evolving diversity of cadherin structures is serving dual and important functions: specific cell adhesion and intricate cell signaling.     

Structure and evolution of the lipase superfamily.

The lipase superfamily includes three vertebrate and three invertebrate (dipteran) proteins that show significant amino acid sequence similarity to one another. The vertebrate proteins are lipoprotein lipase (LPL), hepatic lipase (HL), and pancreatic lipase (PL). The dipteran proteins are Drosophila yolk proteins 1, 2, and 3. We review the relationships among these proteins that have been established according to gene structural relatedness and introduce our findings on the phylogenetic relationships, distance relationships, and evolutionary history of the lipase gene superfamily. Drosophila yolk proteins contain a 104 amino acid residue segment that is conserved with respect to the lipases. We have used the yolk proteins as an outgroup to root a phylogeny of the lipase family. Our phylogenetic reconstruction suggests that ancestral PL diverged earlier than HL and LPL, which share a more recent root. Human and bovine LPL are shown to be more closely related to murine LPL than to guinea pig LPL. A comparison of the distance (a measure of the number of substitutions between sequences) between mammalian and avian LPL reveals that guinea pig LPL has the largest distance from the other mammals. Human, rodent, and rabbit HL show marked divergence from one another, although they have similar relative rates of amino acid substitution when compared to human LPL as an outgroup. Human and porcine PL are not as divergent as human and rat HL, suggesting that PL is more conserved than HL. However, canine PL demonstrates an unusually rapid rate of substitution with respect to the other pancreatic lipases. The lipases share several structurally conserved features. One highly conserved sequence (Gly-Xaa-Ser-Xaa-Gly) contains the active site serine. This feature, which agrees with that found in serine esterases and proteases, is found within the entire spectrum of lipases, including the evolutionarily unrelated prokaryotic lipases. We review the location and possible activity of putative lipid binding domains. We have constructed a conservation index (CI) to display conserved structural features within the lipase gene family, a CI of 1.0 signifying perfect conservation. We have found a correlation between a high CI and the position of conserved functional structures. The putative lipid-binding domains of LPL and HL, the disulfide-bridging cysteine residues, catalytic residues, and N-linked glycosylation sites of LPL, HL, and PL all lie within regions having a CI of 0.8 or higher. A number of amino acid substitutions have been identified in familial hyperchylomicronemia which result in loss of LPL function.(ABSTRACT TRUNCATED AT 400 WORDS)     

Molecular evolution of the arthropod hemocyanin superfamily

Arthropod hemocyanins are members of a protein superfamily that also comprises the arthropod phenoloxidases (tyrosinases), crustacean pseudohemocyanins (cryptocyanins), and insect storage hexamerins. The evolution of these proteins was inferred by neighbor-joining, maximum-parsimony, and maximum-likelihood methods. Monte Carlo shuffling approaches provided evidence against a discernible relationship of the arthropod hemocyanin superfamily and molluscan hemocyanins or nonarthropodan tyrosinases. Within the arthropod hemocyanin superfamily, the phenoloxidase probably emerged early in the (eu-)arthropod stemline and thus form the most likely outgroup. The respiratory hemocyanins evolved from these enzymes before the radiation of the extant euarthropodan subphyla. Due to different functional constraints, replacement rates greatly vary between the clades. Divergence times were thus estimated assuming local molecular clocks using several substitution models. The results were consistent and indicated the separation of the cheliceratan and crustacean hemocyanins close to 600 MYA. The different subunit types of the multihexameric cheliceratan hemocyanin have a rather conservative structure and diversified in the arachnidan stemline between 550 and 450 MYA. By contrast, the separation of the crustacean (malacostracan) hemocyanin subunits probably occurred only about 200 MYA. The nonrespiratory pseudohemocyanins evolved within the Decapoda about 215 MYA. The insect hemocyanins and storage hexamerins emerged independently from the crustacean hemocyanins. The time of divergence of the insect proteins from the malacostracan hemocyanins was estimated to be about 430-440 MYA, providing support for the notion that the Hexapoda evolved from the same crustacean lineage as the Malacostraca.     

核糖核酸酶基因超家族分子进化

核糖核酸酶基因(Ribonuclease A, RNASE A)超家族是进化生物学中研究新基因起源及新功能演变的重要模式系统之一。RNASE A超家族中的很多成员表现出基因复制的进化模式, 而且在适应性(正)选择的驱动下, 发生了功能分化。文章综述了RNASE A超家族成员在不同动物类群中进化模式的研究进展, 包括近年来越来越多在基因组水平上开展的相关研究, 显示该基因超家族可能具有比人们以往认识的更为复杂的基因进化模式。随着越来越多动物基因组数据的产生, 对更多动物代表类群进行RNASE A超家族研究, 将有望揭示新的进化机制和功能分化, 为系统认识动物适应进化的遗传机制奠定基础。     

Structural evolution of the protein kinase-like superfamily

The protein kinase family is large and important, but it is only one family in a larger superfamily of homologous kinases that phosphorylate a variety of substrates and play important roles in all three superkingdoms of life. We used a carefully constructed structural alignment of selected kinases as the basis for a study of the structural evolution of the protein kinase-like superfamily. The comparison of structures revealed a "universal core" domain consisting only of regions required for ATP binding and the phosphotransfer reaction. Remarkably, even within the universal core some kinase structures display notable changes, while still retaining essential activity. Hence, the protein kinase-like superfamily has undergone substantial structural and sequence revision over long evolutionary timescales. We constructed a phylogenetic tree for the superfamily using a novel approach that allowed for the combination of sequence and structure information into a unified quantitative analysis. When considered against the backdrop of species distribution and other metrics, our tree provides a compelling scenario for the development of the various kinase families from a shared common ancestor. We propose that most of the so-called "atypical kinases" are not intermittently derived from protein kinases, but rather diverged early in evolution to form a distinct phyletic group. Within the atypical kinases, the aminoglycoside and choline kinase families appear to share the closest relationship. These two families in turn appear to be the most closely related to the protein kinase family. In addition, our analysis suggests that the actin-fragmin kinase, an atypical protein kinase, is more closely related to the phosphoinositide-3 kinase family than to the protein kinase family. The two most divergent families, alpha-kinases and phosphatidylinositol phosphate kinases (PIPKs), appear to have distinct evolutionary histories. While the PIPKs probably have an evolutionary relationship with the rest of the kinase superfamily, the relationship appears to be very distant (and perhaps indirect). Conversely, the alpha-kinases appear to be an exception to the scenario of early divergence for the atypical kinases: they apparently arose relatively recently in eukaryotes. We present possible scenarios for the derivation of the alpha-kinases from an extant kinase fold.     

Structure and function evolution in the superfamily of globins

The superfamily of globins has emerged some 4000 Myr from a common ancestor, which was among the basic protein components required for life. Globins are present in the three kingdoms of life. From a structure point of view, these molecules are defined by the presence of a characteristic protein fold, rich in α-helix, surrounding a heme group. Depending on the species or organs, they may be physiologically active as monomers, tetramers or large size polymers. Their function varies from the classical reversible binding of oxygen for transport and storage to cytoprotection against reactive oxygen species, NO scavenging, signaling in oxygen dependent metabolic pathways, or possibly other specific properties involving ligand or electron transfer. All these aspects are discussed in this review. To cite this article: H. Wajcman et al., C. R. Biologies 332 (2009).     

节肢动物血蓝蛋白家族的组成与演化

血蓝蛋白是动物界的三类呼吸功能蛋白之一,目前仅发现于节肢动物和软体动物等少数动物类群中。不同亚型的血蓝蛋白有不同的理化性质和序列,但均结合氧分子,并以六聚体,甚至更复杂的聚合体结构存在。血蓝蛋白与酚氧化酶、拟血蓝蛋白、昆虫储存蛋白以及昆虫储存蛋白受体等结构类似、进化上近缘的分子共同组成了血蓝蛋白超家族。该文主要介绍了血蓝蛋白家族成员在节肢动物四大类群（螯肢动物、多足动物、甲壳动物和六足动物）中已知的分布、结构和功能,并重点综述了血蓝蛋白家族成员在节肢动物系统演化研究中发挥的独特而有效的作用,进一步强调了在更多节肢动物类群中研究血蓝蛋白家族的功能和演化的重要性。     

Structure and evolution of the hAT transposon superfamily.

The maize transposon Activator (Ac) was the first mobile DNA element to be discovered. Since then, other elements were found that share similarity to Ac, suggesting that it belongs to a transposon superfamily named hAT after hobo from Drosophila, Ac from maize, and Tam3 from snapdragon. We addressed the structure and evolution of hAT elements by developing new tools for transposon mining and searching the public sequence databases for the hallmarks of hAT elements, namely the transposase and short terminal inverted repeats (TIRs) flanked by 8-bp host duplications. We found 147 hAT-related sequences in plants, animals, and fungi. Six conserved blocks could be identified in the transposase of most hAT elements. A total of 41 hAT sequences were flanked by TIRs and 8-bp host duplications and, out of these, 34 sequences had TIRs similar to the consensus determined in this work, suggesting that they are active or recently active transposons. Phylogenetic analysis and clustering of hAT sequences suggest that the hAT superfamily is very ancient, probably predating the plant-fungi-animal separation, and that, unlike previously proposed, there is no evidence that horizontal gene transfer was involved in the evolution of hAT elements.     

Identification and domain mapping of Dictyostelium discoideum type-1 protein phosphatase inhibitor-2

The protein phosphatase type-1 catalytic subunit (PP1c) does not exist freely in the cell and its activity must be very strictly controlled. Several protein inhibitors of PP1c have been described including the classical mammalian inhibitor-1 (I-1) and inhibitor-2 (I-2). Association of these inhibitors with PP1c appears to involve multiple contacts and in the case of I-2 no less than five I-2 interaction subdomains have been proposed. In this report, we provide both in vitro and in vivo evidence that the Dictyostelium discoideum genome encodes a protein (DdI-2) that is an ortholog of mammalian I-2, being the first PP1c interacting protein characterized in this social amoeba. Despite the low overall sequence similarity of DdI-2 with other I-2 sequences and its long N-terminal extension, the five PP1c interaction motifs proposed for mammalian I-2 are reasonably conserved in the Dictyostelium ortholog. We demonstrate that DdI-2 interacts with and inhibits D. discoideum PP1c (DdPP1c), which we have previously characterized. Moreover, using yeast two-hybrid assays we show that a stable interaction of DdI-2 with DdPP1c requires multiple contacts.     

gammaN-crystallin and the evolution of the betagamma-crystallin superfamily in vertebrates

The beta and gamma crystallins are evolutionarily related families of proteins that make up a large part of the refractive structure of the vertebrate eye lens. Each family has a distinctive gene structure that reflects a history of successive gene duplications. A survey of gamma-crystallins expressed in mammal, reptile, bird and fish species (particularly in the zebrafish, Danio rerio) has led to the discovery of gammaN-crystallin, an evolutionary bridge between the beta and gamma families. In all species examined, gammaN-crystallins have a hybrid gene structure, half beta and half gamma, and thus appear to be the 'missing link' between the beta and gamma crystallin lineages. Overall, there are four major classes of gamma-crystallin: the terrestrial group (including mammalian gammaA-F); the aquatic group (the fish gammaM-crystallins); the gammaS group; and the novel gammaN group. Like the evolutionarily ancient beta-crystallins (but unlike the terrestrial gammaA-F and aquatic gammaM groups), both the gammaS and gammaN crystallins form distinct clades with members in fish, reptiles, birds and mammals. In rodents, gammaN is expressed in nuclear fibers of the lens and, perhaps hinting at an ancestral role for the gamma-crystallins, also in the retina. Although well conserved throughout vertebrate evolution, gammaN in primates has apparently undergone major changes and possible loss of functional expression.     

The evolution of catalytic residues and enzyme mechanism within the bacterial nucleoside phosphorylase superfamily 1

Nucleoside phosphorylases are essential for the salvage and catabolism of nucleotides in bacteria and other organisms, and members of this enzyme superfamily have been of interest for the development of antimicrobial and cancer therapies. The nucleotide phosphorylase superfamily 1 encompasses a number of different enzymes which share a general superfold and catalytic mechanism, while they differ in the nature of the nucleophiles used and in the nature of characteristic active site residues. Recently, one subfamily, the uridine phosphorylases, has been subdivided into two types which differ with respect to the mechanism of transition state stabilization, as dictated by differences in critical amino acid residues. Little is known about the phylogenetic distribution and relationship of the two different types, as well as the relationship to other NP-1 superfamily members. Here comparative genomic analysis illustrates that UP-1s and UP-2s fall into monophyletic groups and are biased with respect to species representation. UP-1 evolved in Gram negative bacteria, while Gram positive species tend to predominantly contain UP-2. PNP (a sister clade to all UPs) contains both Gram positive and Gram negative species. The findings imply that the nucleoside phosphorylase superfamily 1 evolved through a series of three important duplications, leading to the separate, monophyletic enzyme families, coupled to individual lateral transfer events. Extensive horizontal transfer explains the occurrence of unexpected uridine phosphorylases in some genomes. This study provides a basis for understanding the evolution of uridine and purine nucleoside phosphorylases with respect to DNA/RNA metabolism and with potential utility in the design of antimicrobial and anti-tumor drugs.     

Molecular evolution of the Rab-escort-protein/guanine-nucleotide-dissociation-inhibitor superfamily

Prenylation of Rab GTPases regulating vesicle traffic by Rab geranylgeranyltransferase (RabGGTase) requires a complex formed by the association of newly synthesized Rab proteins with Rab-escort-protein (REP), the choroideremia-gene-product that is mutated in disease, leading to loss of vision. After delivery to the membrane by the REP-Rab complex, subsequent recycling to the cytosol requires the REP-related guanine-nucleotide-dissociation-inhibitor (GDI). Although REP and GDI share common Rab-binding properties, GDI cannot assist in Rab prenylation and REP cannot retrieve Rab proteins from the membranes. We have now isolated REP mutant proteins that are able to partially function as both REP and GDI. These results provide molecular insight into the functional and evolutionary organization of the REP/GDI superfamily.