Detection and Alignment of 3D Domain Swapping Proteins Using Angle-Distance Image-Based Secondary Structural Matching Techniques

This work presents a novel detection method for three-dimensional domain swapping (DS), a mechanism for forming protein quaternary structures that can be visualized as if monomers had “opened” their “closed” structures and exchanged the opened portion to form intertwined oligomers. Since the first report of DS in the mid 1990s, an increasing number of identified cases has led to the postulation that DS might occur in a protein with an unconstrained terminus under appropriate conditions. DS may play important roles in the molecular evolution and functional regulation of proteins and the formation of depositions in Alzheimer's and prion diseases. Moreover, it is promising for designing auto-assembling biomaterials. Despite the increasing interest in DS, related bioinformatics methods are rarely available. Owing to a dramatic conformational difference between the monomeric/closed and oligomeric/open forms, conventional structural comparison methods are inadequate for detecting DS. Hence, there is also a lack of comprehensive datasets for studying DS. Based on angle-distance (A-D) image transformations of secondary structural elements (SSEs), specific patterns within A-D images can be recognized and classified for structural similarities. In this work, a matching algorithm to extract corresponding SSE pairs from A-D images and a novel DS score have been designed and demonstrated to be applicable to the detection of DS relationships. The Matthews correlation coefficient (MCC) and sensitivity of the proposed DS-detecting method were higher than 0.81 even when the sequence identities of the proteins examined were lower than 10%. On average, the alignment percentage and root-mean-square distance (RMSD) computed by the proposed method were 90% and 1.8Å for a set of 1,211 DS-related pairs of proteins. The performances of structural alignments remain high and stable for DS-related homologs with less than 10% sequence identities. In addition, the quality of its hinge loop determination is comparable to that of manual inspection. This method has been implemented as a web-based tool, which requires two protein structures as the input and then the type and/or existence of DS relationships between the input structures are determined according to the A-D image-based structural alignments and the DS score. The proposed method is expected to trigger large-scale studies of this interesting structural phenomenon and facilitate related applications

Structural genomics target selection for the New York consortium on membrane protein structure

The New York Consortium on Membrane Protein Structure (NYCOMPS), a part of the Protein Structure Initiative (PSI) in the USA, has as its mission to establish a high-throughput pipeline for determination of novel integral membrane protein structures. Here we describe our current target selection protocol, which applies structural genomics approaches informed by the collective experience of our team of investigators. We first extract all annotated proteins from our reagent genomes, i.e. the 96 fully sequenced prokaryotic genomes from which we clone DNA. We filter this initial pool of sequences and obtain a list of valid targets. NYCOMPS defines valid targets as those that, among other features, have at least two predicted transmembrane helices, no predicted long disordered regions and, except for community nominated targets, no significant sequence similarity in the predicted transmembrane region to any known protein structure. Proteins that feed our experimental pipeline are selected by defining a protein seed and searching the set of all valid targets for proteins that are likely to have a transmembrane region structurally similar to that of the seed. We require sequence similarity aligning at least half of the predicted transmembrane region of seed and target. Seeds are selected according to their feasibility and/or biological interest, and they include both centrally selected targets and community nominated targets. As of December 2008, over 6,000 targets have been selected and are currently being processed by the experimental pipeline. We discuss how our target list may impact structural coverage of the membrane protein space

Glucocortiocoid Treatment of MCMV Infected Newborn Mice Attenuates CNS Inflammation and Limits Deficits in Cerebellar Development

Infection of the developing fetus with human cytomegalovirus (HCMV) is a major cause of central nervous system disease in infants and children; however, mechanism(s) of disease associated with this intrauterine infection remain poorly understood. Utilizing a mouse model of HCMV infection of the developing CNS, we have shown that peripheral inoculation of newborn mice with murine CMV (MCMV) results in CNS infection and developmental abnormalities that recapitulate key features of the human infection. In this model, animals exhibit decreased granule neuron precursor cell (GNPC) proliferation and altered morphogenesis of the cerebellar cortex. Deficits in cerebellar cortical development are symmetric and global even though infection of the CNS results in a non-necrotizing encephalitis characterized by widely scattered foci of virus-infected cells with mononuclear cell infiltrates. These findings suggested that inflammation induced by MCMV infection could underlie deficits in CNS development. We investigated the contribution of host inflammatory responses to abnormal cerebellar development by modulating inflammatory responses in infected mice with glucocorticoids. Treatment of infected animals with glucocorticoids decreased activation of CNS mononuclear cells and expression of inflammatory cytokines (TNF-α, IFN-β and IFNγ) in the CNS while minimally impacting CNS virus replication. Glucocorticoid treatment also limited morphogenic abnormalities and normalized the expression of developmentally regulated genes within the cerebellum. Importantly, GNPC proliferation deficits were normalized in MCMV infected mice following glucocorticoid treatment. Our findings argue that host inflammatory responses to MCMV infection contribute to deficits in CNS development in MCMV infected mice and suggest that similar mechanisms of disease could be responsible for the abnormal CNS development in human infants infected in-utero with HCMV

The small GTPase Arf1 modulates mitochondrial morphology and function

The small GTPase Arf1 plays critical roles in membrane traffic by initiating the recruitment of coat proteins and by modulating the activity of lipid-modifying enzymes. Here, we report an unexpected but evolutionarily conserved role for Arf1 and the ArfGEF GBF1 at mitochondria. Loss of function of ARF-1 or GBF-1 impaired mitochondrial morphology and activity in Caenorhabditis elegans. Similarly, mitochondrial defects were observed in mammalian and yeast cells. In Saccharomyces cerevisiae, aberrant clusters of the mitofusin Fzo1 accumulated in arf1-11 mutants and were resolved by overexpression of Cdc48, an AAA-ATPase involved in ER and mitochondria-associated degradation processes. Yeast Arf1 co-fractionated with ER and mitochondrial membranes and interacted genetically with the contact site component Gem1. Furthermore, similar mitochondrial abnormalities resulted from knockdown of either GBF-1 or contact site components in worms, suggesting that the role of Arf1 in mitochondrial functioning is linked to ER–mitochondrial contacts. Thus, Arf1 is involved in mito- chondrial homeostasis and dynamics, independent of its role in vesicular traffic