355 research outputs found
Evolutionary diversification of the BetaM interactome acquired through co-option of the ATP1B4 gene in placental mammals
ATP1B4 genes represent a rare instance of orthologous vertebrate gene co-option that radically changed properties of the encoded BetaM proteins, which function as Na, K-ATPase subunits in lower vertebrates and birds. Eutherian BetaM has lost its ancestral function and became a muscle-specific resident of the inner nuclear membrane. Our earlier work implicated BetaM in regulation of gene expression through direct interaction with the transcriptional co-regulator SKIP. To gain insight into evolution of BetaM interactome we performed expanded screening of eutherian and avian cDNA libraries using yeast-two-hybrid and split-ubiquitin systems. The inventory of identified BetaM interactors includes lamina-associated protein LAP-1, myocyte nuclear envelope protein Syne1, BetaM itself, heme oxidases HMOX1 and HMOX2; transcription factor LZIP/CREB3, ERGIC3, PHF3, reticulocalbin-3, and beta-sarcoglycan. No new interactions were found for chicken BetaM and human Na, K-ATPase beta 1, beta 2 and beta 3 isoforms, indicating the uniqueness of eutherian BetaM interactome. Analysis of truncated forms of BetaM indicates that residues 72-98 adjacent to the membrane in nucleoplasmic domain are important for the interaction with SKIP. These findings demonstrate that evolutionary alterations in structural and functional properties of eutherian BetaM proteins are associated with the increase in its interactome complexity
Properties of a cryptic lysyl oxidase from haloarchaeon Haloterrigena turkmenica
Background Lysyl oxidases (LOX) have been extensively studied in mammals, whereas properties and functions of recently found homologues in prokaryotic genomes remain enigmatic. Methods LOX open reading frame was cloned from Haloterrigena turkmenica in an E. coli expression vector. Recombinant Haloterrigena turkmenica lysyl oxidase (HTU-LOX) proteins were purified using metal affinity chromatography under denaturing conditions followed by refolding. Amine oxidase activity has been measured fluorometrically as hydrogen peroxide release coupled with the oxidation of 10-acetyl-3,7-dihydroxyphenoxazine in the presence of horseradish peroxidase. Rabbit polyclonal antibodies were obtained and used in western blotting. Results Cultured H. turkmenica has no detectable amine oxidase activity. HTU-LOX may be expressed in E. coli with a high protein yield. The full-length protein gives no catalytic activity. For this reason, we hypothesized that the hydrophobic N-terminal region may interfere with proper folding and its removal may be beneficial. Indeed, truncated His-tagged HTU-LOX lacking the N-terminal hydrophobic signal peptide purified under denaturing conditions can be successfully refolded into an active enzyme, and a larger N-terminal truncation further increases the amine oxidase activity. Refolding is optimal in the presence of Cu2+ at pH 6.2 and is not sensitive to salt. HTU-LOX is sensitive to LOX inhibitor 3-aminopropionitrile. HTU-LOX deaminates usual substrates of mammalian LOX such as lysine-containing polypeptides and polymers. The major difference between HTU-LOX and mammalian LOX is a relaxed substrate specificity of the former. HTU-LOX readily oxidizes various primary amines including such compounds as taurine and glycine, benzylamine being a poor substrate. Of note, HTU-LOX is also active towards several aminoglycoside antibiotics and polymyxin. Western blotting indicates that epitopes for the anti-HTU-LOX polyclonal antibodies coincide with a high molecular weight protein in H. turkmenica cells. Conclusion H. turkmenica contains a lysyl oxidase gene that was heterologously expressed yielding an active recombinant enzyme with important biochemical features conserved between all known LOXes, for example, the sensitivity to 3-aminopropionitrile. However, the native function in the host appears to be cryptic. Significance This is the first report on some properties of a lysyl oxidase from Archaea and an interesting example of evolution of enzymatic properties after hypothetical horizontal transfers between distant taxa
Underlying Event measurements in pp collisions at and 7 TeV with the ALICE experiment at the LHC
Identification of novel protein-protein interactions pertinent to regulation of mammalian hampin/MSL1 and histone acetylase MYST1/MOF
Two distinct nuclear localization signals in mammalian MSL1 regulate its function
MSL1 protein regulates global histone H4 acetylation at residue K16 in stem and cancer cells, through interaction with KAT8. The functional significance of mammalian MSL1 isoforms, involved in various protein interactions, is poorly understood. We report the identification of a novel nuclear localization signal (NLS), common to all MSL1 isoforms, in addition to previously known bipartite NLS, located in domain PEHE. Isoforms having both NLS localize to sub-nuclear foci where they can target co-chaperone protein TTC4. However, all MSL1 isoforms also have ability to affect H4K16 acetylation. Thus, presence of two NLS in MSL1 protein can mediate activity of KAT8 in vivo. J. Cell. Biochem. 115: 1967-1973, 2014. (c) 2014 Wiley Periodicals, Inc
Identification of a novel gene of the X,K-ATPase β-subunit family that is predominantly expressed in skeletal and heart muscles11The nucleotide sequences reported here have been submitted to the GenBank/EBI DATA Bank with accession numbers AF15383, AF15384, AF15385 and AF15386.
AbstractWe have identified the fifth member of the mammalian X,K-ATPase β-subunit gene family. The human and rat genes are largely expressed in skeletal muscle and at a lower level in heart. The deduced human and rat proteins designated as βmuscle (βm) consist of 357 and 356 amino acid residues, respectively, and exhibit 89% identity. The sequence homology of βm proteins with known Na,K- and H,K-ATPase β-subunits are 30.5–39.4%. Unlike other β-subunits, putative βm proteins have large N-terminal cytoplasmic domains containing long Glu-rich sequences. The data obtained indicate the existence of hitherto unknown X,K-ATPase (most probably Na,K-ATPase) isozymes in muscle cells
βm, a Structural Member of the X,K-ATPase β Subunit Family, Resides in the ER and Does Not Associate with Any Known X,K-ATPase α Subunit â€
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