Phosphorylation of Mycobacterium leprae heat-shock 70 protein at threonine 175 alters its substrate binding characteristics. 1998

P Peake, and N Winter, and W Britton
Centenary Institute for Cancer Medicine and Cell Biology, Locked Bag No. 6, Newtown, NSW 2042, Australia.

We have examined the functional properties including autophosphorylation of the Mycobacterium leprae Hsp70 homologue. Recombinant M. leprae Hsp70 had pH optima for its adenosine triphosphatase and autophosphorylating activities which were near pH 8 and 6, respectively. Both these activities were inhibited by reduced and alkylated bovine pancreatic trypsin inhibitor, but not other tested substrates. Autophosphorylation was augmented by up to 25 mM Ca2+. Using site-directed mutagenesis to construct two Thr-->Ala mutants at positions 175 and 193, and phosphoamino acid analysis, it was shown that Thr175 was the dominant threonine residue autophosphorylated in M. leprae Hsp70. Phosphorylation led to an increased affinity for a model polypeptide substrate, reduced and alkylated bovine albumin. These properties are compared with those of the DnaK protein of Escherichia coli.

UI MeSH Term Description Entries
D007611 Aprotinin A single-chain polypeptide derived from bovine tissues consisting of 58 amino-acid residues. It is an inhibitor of proteolytic enzymes including CHYMOTRYPSIN; KALLIKREIN; PLASMIN; and TRYPSIN. It is used in the treatment of HEMORRHAGE associated with raised plasma concentrations of plasmin. It is also used to reduce blood loss and transfusion requirements in patients at high risk of major blood loss during and following open heart surgery with EXTRACORPOREAL CIRCULATION. (Reynolds JEF(Ed): Martindale: The Extra Pharmacopoeia (electronic version). Micromedex, Inc, Englewood, CO, 1995) BPTI, Basic Pancreatic Trypsin Inhibitor,Basic Pancreatic Trypsin Inhibitor,Bovine Kunitz Pancreatic Trypsin Inhibitor,Kallikrein-Trypsin Inactivator,Kunitz Pancreatic Trypsin Inhibitor,Trypsin Inhibitor, Basic, Pancreatic,Trypsin Inhibitor, Kunitz, Pancreatic,Antilysin,Bovine Pancreatic Trypsin Inhibitor,Contrical,Contrykal,Dilmintal,Iniprol,Kontrikal,Kontrykal,Pulmin,Traskolan,Trasylol,Zymofren,Inactivator, Kallikrein-Trypsin,Kallikrein Trypsin Inactivator
D009154 Mutation Any detectable and heritable change in the genetic material that causes a change in the GENOTYPE and which is transmitted to daughter cells and to succeeding generations. Mutations
D009166 Mycobacterium leprae A species of gram-positive, aerobic bacteria that causes LEPROSY in man. Its organisms are generally arranged in clumps, rounded masses, or in groups of bacilli side by side.
D010766 Phosphorylation The introduction of a phosphoryl group into a compound through the formation of an ester bond between the compound and a phosphorus moiety. Phosphorylations
D010769 Phosphothreonine The phosphoric acid ester of threonine. Used as an identifier in the analysis of peptides, proteins, and enzymes. Threonine Phosphate,Phosphate, Threonine
D011485 Protein Binding The process in which substances, either endogenous or exogenous, bind to proteins, peptides, enzymes, protein precursors, or allied compounds. Specific protein-binding measures are often used as assays in diagnostic assessments. Plasma Protein Binding Capacity,Binding, Protein
D011994 Recombinant Proteins Proteins prepared by recombinant DNA technology. Biosynthetic Protein,Biosynthetic Proteins,DNA Recombinant Proteins,Recombinant Protein,Proteins, Biosynthetic,Proteins, Recombinant DNA,DNA Proteins, Recombinant,Protein, Biosynthetic,Protein, Recombinant,Proteins, DNA Recombinant,Proteins, Recombinant,Recombinant DNA Proteins,Recombinant Proteins, DNA
D002118 Calcium A basic element found in nearly all tissues. It is a member of the alkaline earth family of metals with the atomic symbol Ca, atomic number 20, and atomic weight 40. Calcium is the most abundant mineral in the body and combines with phosphorus to form calcium phosphate in the bones and teeth. It is essential for the normal functioning of nerves and muscles and plays a role in blood coagulation (as factor IV) and in many enzymatic processes. Coagulation Factor IV,Factor IV,Blood Coagulation Factor IV,Calcium-40,Calcium 40,Factor IV, Coagulation
D006863 Hydrogen-Ion Concentration The normality of a solution with respect to HYDROGEN ions; H+. It is related to acidity measurements in most cases by pH pH,Concentration, Hydrogen-Ion,Concentrations, Hydrogen-Ion,Hydrogen Ion Concentration,Hydrogen-Ion Concentrations
D000251 Adenosine Triphosphatases A group of enzymes which catalyze the hydrolysis of ATP. The hydrolysis reaction is usually coupled with another function such as transporting Ca(2+) across a membrane. These enzymes may be dependent on Ca(2+), Mg(2+), anions, H+, or DNA. ATPases,Adenosinetriphosphatase,ATPase,ATPase, DNA-Dependent,Adenosine Triphosphatase,DNA-Dependent ATPase,DNA-Dependent Adenosinetriphosphatases,ATPase, DNA Dependent,Adenosinetriphosphatases, DNA-Dependent,DNA Dependent ATPase,DNA Dependent Adenosinetriphosphatases,Triphosphatase, Adenosine

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