Mutational analysis of βCOP (Sec26p) identifies an appendage domain critical for function

Background  

The appendage domain of the γCOP subunit of the COPI vesicle coat bears a striking structural resemblance to adaptin-family appendages despite limited primary sequence homology. Both the γCOP appendage domain and an equivalent region on βCOP contain the FxxxW motif; the conservation of this motif suggested the existence of a functional appendage domain in βCOP.     

The BACK domain in BTB-kelch proteins

A novel conserved motif--the BACK (for BTB and C-terminal Kelch) domain--is found in the majority of proteins that contain both the BTB domain and kelch repeats. Many kelch-repeat proteins are involved in organization of the cytoskeleton via interaction with actin and intermediate filaments, whereas BTB domains have multiple cellular roles, including recruitment to E3 ubiquitin ligase complexes. The identification of the BACK domain in BTB and kelch proteins, and its high conservation across metazoan genomes, suggest an important function for this domain with a possible role in substrate orientation in Cullin3-based E3 ligase complexes.     

Sequence and structural analysis of the Asp-box motif and Asp-box beta-propellers; a widespread propeller-type characteristic of the Vps10 domain family and several glycoside hydrolase families

Background  

The Asp-box is a short sequence and structure motif that folds as a well-defined β-hairpin. It is present in different folds, but occurs most prominently as repeats in β-propellers. Asp-box β-propellers are known to be characteristically irregular and to occur in many medically important proteins, most of which are glycosidase enzymes, but they are otherwise not well characterized and are only rarely treated as a distinct β-propeller family. We have analyzed the sequence, structure, function and occurrence of the Asp-box and s-Asp-box -a related shorter variant, and provide a comprehensive classification and computational analysis of the Asp-box β-propeller family.     

Thiol dioxygenases: unique families of cupin proteins

Proteins in the cupin superfamily have a wide range of biological functions in archaea, bacteria and eukaryotes. Although proteins in the cupin superfamily show very low overall sequence similarity, they all contain two short but partially conserved cupin sequence motifs separated by a less conserved intermotif region that varies both in length and amino acid sequence. Furthermore, these proteins all share a common architecture described as a six-stranded β-barrel core, and this canonical cupin or “jelly roll” β-barrel is formed with cupin motif 1, the intermotif region, and cupin motif 2 each forming two of the core six β-strands in the folded protein structure. The recently obtained crystal structures of cysteine dioxygenase (CDO), with contains conserved cupin motifs, show that it has the predicted canonical cupin β-barrel fold. Although there had been no reports of CDO activity in prokaryotes, we identified a number of bacterial cupin proteins of unknown function that share low similarity with mammalian CDO and that conserve many residues in the active-site pocket of CDO. Putative bacterial CDOs predicted to have CDO activity were shown to have similar substrate specificity and kinetic parameters as eukaryotic CDOs. Information gleaned from crystal structures of mammalian CDO along with sequence information for homologs shown to have CDO activity facilitated the identification of a CDO family fingerprint motif. One key feature of the CDO fingerprint motif is that the canonical metal-binding glutamate residue in cupin motif 1 is replaced by a cysteine (in mammalian CDOs) or by a glycine (bacterial CDOs). The recent report that some putative bacterial CDO homologs are actually 3-mercaptopropionate dioxygenases suggests that the CDO family may include proteins with specificities for other thiol substrates. A paralog of CDO in mammals was also identified and shown to be the other mammalian thiol dioxygenase, cysteamine dioxygenase (ADO). A tentative fingerprint motif for ADOs, or DUF1637 family members, is proposed. In ADOs, the conserved glutamate residue in cupin motif 1 is replaced by either glycine or valine. Both ADOs and CDOs appear to represent unique clades within the cupin superfamily.     

The kelch repeat superfamily of proteins: propellers of cell function

The kelch motif was discovered as a sixfold tandem element in the sequence of the Drosophila kelch ORF1 protein. The repeated kelch motifs predict a conserved tertiary structure, a beta-propeller. This module appears in many different polypeptide contexts and contains multiple potential protein-protein contact sites. Members of this growing superfamily are present throughout the cell and extracellularly and have diverse activities. In this review, we discuss current information concerning the structural organization of kelch repeat proteins, their biological roles and the molecular basis of their action.     

Sequencing and comparative analysis of fugu protocadherin clusters reveal diversity of protocadherin genes among teleosts

Background  

The synaptic cell adhesion molecules, protocadherins, are a vertebrate innovation that accompanied the emergence of the neural tube and the elaborate central nervous system. In mammals, the protocadherins are encoded by three closely-linked clusters (α, β and γ) of tandem genes and are hypothesized to provide a molecular code for specifying the remarkably-diverse neural connections in the central nervous system. Like mammals, the coelacanth, a lobe-finned fish, contains a single protocadherin locus, also arranged into α, β and γ clusters. Zebrafish, however, possesses two protocadherin loci that contain more than twice the number of genes as the coelacanth, but arranged only into α and γ clusters. To gain further insight into the evolutionary history of protocadherin clusters, we have sequenced and analyzed protocadherin clusters from the compact genome of the pufferfish, Fugu rubripes.     

Structural characterization of plant defensin protein superfamily

Plant defensins represent a major innate immune protein superfamily with strong inhibitory effects on infectious diseases of humans, antifungal/antibacterial activities, proteinase and insect amylase inhibitory activities. They are generally defined by their conserved cysteine scaffold with α-helix and triple strand anti parallel β-sheet connected to the scaffold. With the genome of more plant species being fully sequenced, significant information about newly sequenced defensin proteins has been revealed. In this paper, we identify members of defensin protein families across plant species and use protein-modeling-based structural reconstitution to reveal specific three dimensional hidden features of plant defensins mediating defense responses and other interesting biological activities in plants. Our data revealed that plant defensins are structurally similar to their insect counterparts despite the low amino acid sequence similarity between these two organisms. The molecular and structural relationship among plant defensins and defensins from other species is discussed.     

<Emphasis Type="Italic">Drosophila</Emphasis> Uri,a PP1α binding protein,is essential for viability,maintenance of DNA integrity and normal transcriptional activity

Background  

Protein phosphatase 1 (PP1) is involved in diverse cellular processes, and is targeted to substrates via interaction with many different protein binding partners. PP1 catalytic subunits (PP1c) fall into PP1α and PP1β subfamilies based on sequence analysis, however very few PP1c binding proteins have been demonstrated to discriminate between PP1α and PP1β.     

The origin of bmp16, a novel Bmp2/4 relative, retained in teleost fish genomes

Background  

Whole genome sequences have allowed us to have an overview of the evolution of gene repertoires. The target of the present study, the TGFβ superfamily, contains many genes involved in vertebrate development, and provides an ideal system to explore the relationships between evolution of gene repertoires and that of developmental programs.     

A combination of the F-box motif and kelch repeats defines a large Arabidopsis family of F-box proteins

In the sequences released by the Arabidopsis Genome Initiative (AGI), we have discovered a new large gene family (48 genes as of July 2000). A detailed computational and biochemical analysis of the predicted gene products reveals a novel family of plant F-box proteins, where the amino (N)-terminal F-box motif is followed by four kelch repeats and a characteristic carboxy-terminal domain. F-box proteins are an expanding family of eukaryotic proteins, which have been shown in some cases to be critical for the controlled degradation of cellular regulatory proteins via the ubiquitin pathway. The F-box motif of the At5g48990 gene product, a member of the family, was shown to be functionally active by its ability to mediate the in vitro interaction between At5g48990 and ASK1 proteins. F-box proteins specifically recruit the targets to be ubiquitinated, mainly through protein-protein interaction modules such as WD-40 domains or leucine-rich repeats (LRRs). The kelch repeats of the family described here form a potential protein-protein interaction domain, as molecular modelling of the kelch repeats according to the galactose oxidase crystal structure (the only solved structure containing kelch repeats) predicts a -propeller. The identification of this family of F-box proteins greatly expands the field of plant F-box proteins and suggests that controlled degradation of cellular proteins via the ubiquitin pathway could play a critical role in multiple plant cellular processes.     

pi-Turns: types,systematics and the context of their occurrence in protein structures

Background  

For a proper understanding of protein structure and folding it is important to know if a polypeptide segment adopts a conformation inherent in the sequence or it depends on the context of its flanking secondary structures. Turns of various lengths have been studied and characterized starting from three-residue γ-turn to six-residue π-turn. The Schellman motif occurring at the C-terminal end of α-helices is a classical example of hydrogen bonded π-turn involving residues at (i) and (i+5) positions. Hydrogen bonded and non-hydrogen bonded β- and α-turns have been identified previously; likewise, a systematic characterization of π-turns would provide valuable insight into turn structures.     

A Hidden Markov Model method,capable of predicting and discriminating β-barrel outer membrane proteins

Background  

Integral membrane proteins constitute about 20–30% of all proteins in the fully sequenced genomes. They come in two structural classes, the α-helical and the β-barrel membrane proteins, demonstrating different physicochemical characteristics, structure and localization. While transmembrane segment prediction for the α-helical integral membrane proteins appears to be an easy task nowadays, the same is much more difficult for the β-barrel membrane proteins. We developed a method, based on a Hidden Markov Model, capable of predicting the transmembrane β-strands of the outer membrane proteins of gram-negative bacteria, and discriminating those from water-soluble proteins in large datasets. The model is trained in a discriminative manner, aiming at maximizing the probability of correct predictions rather than the likelihood of the sequences.     

Prediction of "hot spots" of aggregation in disease-linked polypeptides

Background  

The polypeptides involved in amyloidogenesis may be globular proteins with a defined 3D-structure or natively unfolded proteins. The first class includes polypeptides such as β2-microglobulin, lysozyme, transthyretin or the prion protein, whereas β-amyloid peptide, amylin or α-synuclein all belong to the second class. Recent studies suggest that specific regions in the proteins act as "hot spots" driving aggregation. This should be especially relevant for natively unfolded proteins or unfolded states of globular proteins as they lack significant secondary and tertiary structure and specific intra-chain interactions that can mask these aggregation-prone regions. Prediction of such sequence stretches is important since they are potential therapeutic targets.     

Cloning and characterization of a β-1,3-glucan-binding protein from shrimp <Emphasis Type="Italic">Fenneropenaeus chinensis</Emphasis>

The β-1,3-d-glucan binding protein (βGBP), one kind of the pattern recognition proteins (PRPs), was cloned and characterized from the Chinese Shrimp Fenneropenaeus chinensis, and named as FcβGBP-HDL. The results indicated that the full length cDNA of 6713 bp had an open reading frame encoded a polypeptide of 2139 amino acids with two glucanase-like motifs and one RGD motif, while without signal peptide. The calculated molecular mass of mature protein was 240.7 kDa and theoretical pI was 5.95. Sequence comparison of the deduced amino acid sequence of FcβGBP-HDL showed varied identity of 88, 54 and 53% with those of Litopenaeus vannamei βGBP-HDL, Pacifastacus leniusculus βGBP, and Pontastacus leptodactylus βGBP, respectively. qRT-PCR analysis indicated that FcβGBP-HDL was expressed in intestines, hepatopancreas, muscle, gill and hemocytes, and its profile was modified post WSSV challenge. The differential expressions of FcβGBP-HDL in different tissues post WSSV challenge suggested that FcβGBP-HDL might play an important role in shrimp immune and perform differently in different tissues. These data would be helpful to better understand the WSSV-resistance mechanism of farming shrimp.     

Delayed internalization and lack of recycling in a beta<Subscript>2</Subscript>-adrenergic receptor fused to the G protein alpha-subunit

Background  

Chimeric proteins obtained by the fusion of a G protein-coupled receptor (GPCR) sequence to the N-terminus of the G protein α-subunit have been extensively used to investigate several aspects of GPCR signalling. Although both the receptor and the G protein generally maintain a fully functional state in such polypeptides, original observations made using a chimera between the β₂-adrenergic receptor (β₂AR) and Gα_s indicated that the fusion to the α-subunit resulted in a marked reduction of receptor desensitization and down-regulation. To further investigate this phenomenon, we have compared the rates of internalization and recycling between wild-type and Gα_s-fused β₂AR.     

Glycogen synthase kinase 3α and 3β have distinct functions during cardiogenesis of zebrafish embryo

Background  

Glycogen synthase kinase 3 (GSK3) encodes a serine/threonine protein kinase, is known to play roles in many biological processes. Two closely related GSK3 isoforms encoded by distinct genes: GSK3α (51 kDa) and GSK3β (47 kDa). In previously studies, most GSK3 inhibitors are not only inhibiting GSK3, but are also affecting many other kinases. In addition, because of highly similarity in amino acid sequence between GSK3α and GSK3β, making it difficult to identify an inhibitor that can be selective against GSK3α or GSK3β. Thus, it is relatively difficult to address the functions of GSK3 isoforms during embryogenesis. At this study, we attempt to specifically inhibit either GSK3α or GSK3β and uncover the isoform-specific roles that GSK3 plays during cardiogenesis.     

Regulation of genes affecting body size and innate immunity by the DBL-1/BMP-like pathway in <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis>

Background  

Bone morphogenetic proteins (BMPs) are members of the conserved transforming growth factor β (TGFβ superfamily, and play many developmental and homeostatic roles. In C. elegans, a BMP-like pathway, the DBL-1 pathway, controls body size and is involved in innate immunity. How these functions are carried out, though, and what most of the downstream targets of this pathway are, remain unknown.     

Gα<Subscript>16</Subscript> interacts with tetratricopeptide repeat 1 (TPR1) through its β3 region to activate Ras independently of phospholipase Cβ signaling

Background  

G protein-coupled receptors constitute the largest family of cell surface receptors in the mammalian genome. As the core of the G protein signal transduction machinery, the Gα subunits are required to interact with multiple partners. The GTP-bound active state of many Gα subunits can bind a multitude of effectors and regulatory proteins. Yet it remains unclear if the different proteins utilize distinct or common structural motifs on the Gα subunit for binding. Using Gα₁₆ as a model, we asked if its recently discovered adaptor protein tetratricopeptide repeat 1 (TPR1) binds to the same region as its canonical effector, phospholipase Cβ (PLCβ).