@article {505, title = {Deciphering Mode of Action of Functionally Important Regions in the Intrinsically Disordered Paxillin (Residues 1-313) Using Its Interaction with FAT (Focal Adhesion Targeting Domain of Focal Adhesion Kinase). [Protein Technology Core]}, journal = {PLoS One}, volume = {11}, year = {2016}, month = {2016}, pages = {e0150153}, abstract = {

Intrinsically disordered proteins (IDPs) play a major role in various cellular functions ranging from transcription to cell migration. Mutations/modifications in such IDPs are shown to be associated with various diseases. Current strategies to study the mode of action and regulatory mechanisms of disordered proteins at the structural level are time consuming and challenging. Therefore, using simple and swift strategies for identifying functionally important regions in unstructured segments and understanding their underlying mechanisms is critical for many applications. Here we propose a simple strategy that employs dissection of human paxillin (residues 1-313) that comprises intrinsically disordered regions, followed by its interaction study using FAT (Focal adhesion targeting domain of focal adhesion kinase) as its binding partner to retrace structural behavior. Our findings show that the paxillin interaction with FAT exhibits a masking and unmasking effect by a putative intra-molecular regulatory region. This phenomenon suggests how cancer associated mutations in paxillin affect its interactions with Focal Adhesion Kinase (FAK). The strategy could be used to decipher the mode of regulations and identify functionally relevant constructs for other studies.

}, keywords = {Focal Adhesion Protein-Tyrosine Kinases, Focal Adhesions, Humans, Intrinsically Disordered Proteins, Models, Molecular, Paxillin, Peptide Fragments, Protein Binding, Protein Structure, Tertiary}, issn = {1932-6203}, doi = {10.1371/journal.pone.0150153}, author = {Neerathilingam, Muniasamy and Bairy, Sneha G and Mysore, Sumukh} } @article {513, title = {L-Plastin S-glutathionylation promotes reduced binding to β-actin and affects neutrophil functions. (Mass Spectrometry)}, journal = {Free Radic Biol Med}, volume = {86}, year = {2015}, month = {2015 Sep}, pages = {1-15}, abstract = {

Posttranslational modifications (PTMs) of cytoskeleton proteins due to oxidative stress associated with several pathological conditions often lead to alterations in cell function. The current study evaluates the effect of nitric oxide (DETA-NO)-induced oxidative stress-related S-glutathionylation of cytoskeleton proteins in human PMNs. By using in vitro and genetic approaches, we showed that S-glutathionylation of L-plastin (LPL) and β-actin promotes reduced chemotaxis, polarization, bactericidal activity, and phagocytosis. We identified Cys-206, Cys-283, and Cys-460as S-thiolated residues in the β-actin-binding domain of LPL, where cys-460 had the maximum score. Site-directed mutagenesis of LPL Cys-460 further confirmed the role in the redox regulation of LPL. S-Thiolation diminished binding as well as the bundling activity of LPL. The presence of S-thiolated LPL was detected in neutrophils from both diabetic patients and db/db mice with impaired PMN functions. Thus, enhanced nitroxidative stress may results in LPL S-glutathionylation leading to impaired chemotaxis, polarization, and bactericidal activity of human PMNs, providing a mechanistic basis for their impaired functions in diabetes mellitus.

}, keywords = {Actins, Adult, Amino Acid Sequence, Animals, Case-Control Studies, Cell Polarity, Chemotaxis, Diabetes Mellitus, Female, Glutathione, HEK293 Cells, Humans, Male, Mice, Inbred C57BL, Mice, Obese, Microfilament Proteins, Middle Aged, Molecular Sequence Data, Neutrophils, Nitric Oxide, Oxidative Stress, Protein Binding, Protein Processing, Post-Translational, Young Adult}, issn = {1873-4596}, doi = {10.1016/j.freeradbiomed.2015.04.008}, author = {Dubey, Megha and Singh, Abhishek K and Awasthi, Deepika and Nagarkoti, Sheela and Kumar, Sachin and Ali, Wahid and Chandra, Tulika and Kumar, Vikas and Barthwal, Manoj K and Jagavelu, Kumaravelu and S{\'a}nchez-G{\'o}mez, Francisco J and Lamas, Santiago and Dikshit, Madhu} } @article {484, title = {Genomic analysis reveals epistatic silencing of "expensive" genes in Escherichia coli K-12. [Next Generation Genomics facility]}, journal = {Mol Biosyst}, volume = {9}, year = {2013}, month = {2013 Aug}, pages = {2021-33}, abstract = {

A barrier for horizontal gene transfer is high gene expression, which is metabolically expensive. Silencing of horizontally-acquired genes in the bacterium Escherichia coli is caused by the global transcriptional repressor H-NS. The activity of H-NS is enhanced or diminished by other proteins including its homologue StpA, and Hha and YdgT. The interconnections of H-NS with these regulators and their role in silencing gene expression in E. coli are not well understood on a genomic scale. In this study, we use transcriptome sequencing to show that there is a bi-layered gene silencing system - involving the homologous H-NS and StpA - operating on horizontally-acquired genes among others. We show that H-NS-repressed genes belong to two types, termed "epistatic" and "unilateral". In the absence of H-NS, the expression of "epistatically controlled genes" is repressed by StpA, whereas that of "unilaterally controlled genes" is not. Epistatic genes show a higher tendency to be non-essential and recently acquired, when compared to unilateral genes. Epistatic genes reach much higher expression levels than unilateral genes in the absence of the silencing system. Finally, epistatic genes contain more high affinity H-NS binding motifs than unilateral genes. Therefore, both the DNA binding sites of H-NS as well as the function of StpA as a backup system might be selected for silencing highly transcribable genes.

}, keywords = {Binding Sites, DNA-Binding Proteins, Epistasis, Genetic, Escherichia coli K12, Escherichia coli Proteins, Fimbriae Proteins, Gene Expression Regulation, Bacterial, Gene Silencing, Gene Transfer, Horizontal, Genome, Bacterial, Molecular Chaperones, Protein Binding, Repressor Proteins, Sequence Analysis, DNA, Transcription, Genetic, Transcriptome}, issn = {1742-2051}, doi = {10.1039/c3mb70035f}, author = {Srinivasan, Rajalakshmi and Chandraprakash, Deepti and Krishnamurthi, Revathy and Singh, Parul and Scolari, Vittore F and Krishna, Sandeep and Seshasayee, Aswin Sai Narain} }