title: Loss of protection by nucleotides against proteolysis and thiol modification in the isolated alpha-subunit from F1 ATPase of Escherichia coli mutant uncA401.
abstract: Binding of nucleotides to the high-affinity site of the isolated alpha subunit of normal Escherichia coli F1 adenosine triphosphatase (ATPase) results in partial protection against digestion by trypsin [Senda, Kanazawa, Tsuchiya & Futai (1983) Arch. Biochem. Biophys. 220, 398-440]. In contrast, the isolated alpha subunit from the defective ATPase of the E. coli uncA401 mutant (strain AN120) is cleaved by trypsin to peptides of less than 8000 Da in the presence of ADP or ATP (2.5 microM-110 mM). The nucleotide-dependent accessibility of thiol groups of the isolated alpha subunit was also studied. Two out of four thiol groups of the alpha subunit from normal ATPase are labelled by fluorescent maleimides or iodoacetates, but in the presence of ADP or ATP (0.14-1.2 mM), reaction of thiol groups with these labels is almost absent. Mutant alpha subunit, however, is labelled by these reagents at all four thiol groups in the presence or absence of ADP or ATP (1 mM). These results suggest that the mutation in the ATPase of strain AN120 leads either to the loss of the high-affinity nucleotide-binding site or affects transmission of allosteric changes that occur on binding of nucleotide to the isolated alpha subunit.
mesh_terms: D000244:Adenosine Diphosphate / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D009154:Mutation; D009711:Nucleotides / Q000378:metabolism*; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000378:metabolism; D014357:Trypsin / Q000494:pharmacology
chemical_list: D009711:Nucleotides; D013438:Sulfhydryl Compounds; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D014357:Trypsin; D006180:Proton-Translocating ATPases