Synthesis and Biochemical Characterization of an Analogue of CheY-Phosphate, a Signal Transduction Protein in Bacterial Chemotaxis†

Biochemistry ◽  
1998 ◽  
Vol 37 (39) ◽  
pp. 13674-13680 ◽  
Author(s):  
Christopher J. Halkides ◽  
Xiangyang Zhu ◽  
Dennis P. Phillion ◽  
Philip Matsumura ◽  
Frederick W. Dahlquist
Keyword(s):  
Signal Transduction ◽  
Biochemical Characterization ◽  
Bacterial Chemotaxis ◽  
Signal Transduction Protein

scholarly journals Cloning and biochemical characterization of the cyclophilin homologues from the free-living nematode Caenorhabditis elegans

1996 ◽  
Vol 317 (1) ◽  
pp. 179-185 ◽  
Author(s):  
Antony P. PAGE ◽  
Kenneth MacNIVEN ◽  
Michael O. HENGARTNER
Keyword(s):  
Signal Transduction ◽  
Protein Folding ◽  
Caenorhabditis Elegans ◽  
Biochemical Characterization ◽  
Single Species ◽  
Immunosuppressive Agent ◽  
Specific Substrate ◽  
Nematode Caenorhabditis Elegans ◽  
Isomerase Activity

Cyclosporin A (CsA) is the most widely used immunosuppressive agent, whose properties are exerted via an interaction with cyclophilin, resulting in down-regulation of signal-transduction events in the T-cell. Cyclophilin is identical with peptidylprolyl cis–trans isomerase (PPI; EC 5.2.1.8), an enzyme which catalyses the isomerization between the two proline conformations in proteins, thereby acting as a catalyst in protein-folding events. Several reports indicate that CsA has potent anti-parasitic activity, effective against both protozoan and helminth species. In order to understand the various biological roles that cyclophilins play we have initiated a study of these proteins in the genetically tractable nematode Caenorhabditis elegans. Here we describe the cloning and characterization of 11 cyclophilin genes (cyp-1 to -11) derived from this nematode; this is currently the greatest number of isoforms described in a single species. Southern blotting and physical mapping indicated that these genes are dispersed throughout the nematode genome. A high degree of conservation exists between several isoforms, which also share characteristics with the ubiquitous isoforms previously described. The remaining isoforms are divergent, having altered CsA-binding domains and additional non-cyclophilin domains, which may impart compartmental specificity. Ten of these isoforms have been expressed in Escherichia coli, and the resultant fusion proteins have been examined biochemically for PPI activity, which they all possess. Isomerase activity is highest in the conserved and lowest in divergent isoforms, perhaps indicating a more specific substrate for the latter. Analysis of the C. elegans cyp genes will provide answers as to the roles played by cyclophilins in protein folding and signal transduction.

scholarly journals Characterization of atrial-natriuretic-factor-receptor-coupled membrane guanylate cyclase from rat and mouse testes

1988 ◽  
Vol 251 (1) ◽  
pp. 301-304 ◽  
Author(s):  
R B Marala ◽  
R K Sharma
Keyword(s):  
Signal Transduction ◽  
Guanylate Cyclase ◽  
Adrenocortical Carcinoma ◽  
Atrial Natriuretic Factor ◽  
Biochemical Characterization ◽  
Membrane Guanylate Cyclase ◽  
Natriuretic Factor ◽  
Previous Demonstration ◽  
Atrial Natriuretic

Studies with isolated adrenal cells and mouse testicular cells have supported a mediatory role of cyclic GMP in ANF (atrial natriuretic factor)-dependent steroidogenic signal transduction. This concept has been strengthened by the purification and biochemical characterization of a 180 kDa protein, which appears to contain both ANF receptor and guanylate cyclase activities, from rat adrenocortical carcinoma cells. Utilizing the antibody to 180 kDa membrane guanylate cyclase as a probe, we now demonstrate the direct presence of ANF-dependent membrane guanylate cyclase in mouse and rat testes. The antibody blocks the ANF-dependent guanylate cyclase activity in isolated membranes, and Western-blot analysis of the partially purified enzyme reveals a single 180 kDa protein. The presence of this enzyme in mouse and rat testes, together with its previous demonstration in rat adrenocortical carcinoma, represent an important potential biochemical role for this enzyme in receptor-mediated steroidogenic signal transduction.

scholarly journals An HD domain phosphohydrolase active site tailored for oxetanocin-A biosynthesis

2016 ◽  
Vol 113 (48) ◽  
pp. 13750-13755 ◽  
Author(s):  
Jennifer Bridwell-Rabb ◽  
Gyunghoon Kang ◽  
Aoshu Zhong ◽  
Hung-wen Liu ◽  
Catherine L. Drennan
Keyword(s):  
Signal Transduction ◽  
Crystal Structures ◽  
Bacillus Megaterium ◽  
Active Site ◽  
Inorganic Phosphate ◽  
Biochemical Characterization ◽  
Membered Ring ◽  
Nucleotide Metabolism ◽  
X Ray

HD domain phosphohydrolase enzymes are characterized by a conserved set of histidine and aspartate residues that coordinate an active site metallocenter. Despite the important roles these enzymes play in nucleotide metabolism and signal transduction, few have been both biochemically and structurally characterized. Here, we present X-ray crystal structures and biochemical characterization of theBacillus megateriumHD domain phosphohydrolase OxsA, involved in the biosynthesis of the antitumor, antiviral, and antibacterial compound oxetanocin-A. These studies reveal a previously uncharacterized reaction for this family; OxsA catalyzes the conversion of a triphosphorylated compound into a nucleoside, releasing one molecule of inorganic phosphate at a time. Remarkably, this functionality is a result of the OxsA active site, which based on structural and kinetic analyses has been tailored to bind the small, four-membered ring of oxetanocin-A over larger substrates. Furthermore, our OxsA structures show an active site that switches from a dinuclear to a mononuclear metal center as phosphates are eliminated from substrate.

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