We have determined the sequence of the E. coli gyrB gene, using a new sequencing approach in which transposition from a mini-Mu plasmid into the DNA provides random start points for dideoxynucleotide sequence analysis. The gyrB sequence corresponds to a protein 804 amino acids long; a previously isolated protein fragment with partial enzymatic activity has been identified as the C-terminal half-molecule. A plausible terminator of gyrB transcription is located just beyond the structural gene.
In Klebsiella aerogenes, arylsulfatase synthesis was repressed by inorganic sulfate, sulfite, sulfide, thiosulfate, and cysteine, but not by methionine under normal growth conditions. We isolated cysteine-requiring mutants (Cys-), and mutants (AtsS-, AtsR-) in which the regulation of arylsulfatase synthesis was altered. In the cysteine auxotroph, enzyme synthesis was also repressed by inorganic sulfate or cysteine. Kinetic studies on mutants of the cysteine auxotroph showed that inorganic sulfate repressed arylsulfatase synthesis and that this was not due to cysteine formed by reduction of sulfate. Arylsulfatase synthesis in the AtsSmutant was not repressed by inorganic sulfate but was repressed by cysteine. This mutant strain had a normal level of inorganic sulfate transport. Another mutant strain, defective in the inorganic sulfate transport system, synthesized arylsulfatase in the presence of inorganic sulfate but not in the presence of cysteine. The AtsSmutant could synthesize the enzyme in the presence of inorganic sulfate but not cysteine. The AtsRmutant could synthesize the enzyme in the presence of either inorganic sulfate or cysteine. These results suggest that there are two independent functional corepressors of arylsulfatase synthesis in K. aerogenes. Harada and Spencer (8) showed that, in many fungi, arylsulfatase synthesis is inhibited by the presence of sulfur compounds, such as sulfate, sulfite, thiosulfate, and cysteine, which are thought to be direct intermediates in the assimilation of sulfate, but that it is not inhibited by compounds such as methionine, taurine, and cysteate, which are not direct intermediates. A similar division of sulfur sources was observed in the arylsulfatase synthesis of Aerobacter aerogenes (9). The effects of various sulfur compounds in repressing arylsulfatase synthesis has been thought to be due to conversion of these compounds to a single corepressor compound: inorganic sulfate in the case of fungi and cysteine in the case of A. aerogenes. The work described in this paper indicates that there may be two independent functional corepressors of arylsulfatase synthesis in Klebsiella aerogenes W70. MATERIALS AND METHODS Bacterial strains and growth conditions. The strains employed in this work are listed in Table 1. They are derivatives of K. aerogenes W70 described by MacPhee et al. (11). Organisms were cultured as described previously (1), except that 50 1sg of L-leucine, L-cysteine, or both were added per mg when required, and cells were grown at 37 C.
It was shown that at least four genes are specifically responsible for arylsulfatase synthesis inKlebsiella aerogenes. Mutations at chromosome site atsA result in enzymatically inactive arylsulfatase. Mutants showing constitutive synthesis of arylsulfatase (atsR) were isolated by using inorganic sulfate or cysteine as the sulfur source. Another mutation in which repression of arylsulfatase by inorganic sulfate or cysteine could not be relieved by tyramine was determined by genetic analysis to be on the tyramine oxidase gene (tyn). This site was distinguished from the atsC mutation site, which is probably concerned with the action or synthesis of corepressors of arylsulfatase synthesis. Genetic analysis with transducing phage PW52 showed that the order of mutation sites was atsC-atsR-atsA-tynA-tynB. On the basis of these results and previous physiological findings, we propose a new model for regulation of arylsulfatase synthesis.
Studies were made on the effect of tyramine on arylsulfatase synthesis in mutants of Aerobacter aerogenes ATCC 9621 deficient in enzymes involved in tyramine degradation. As shown previously, some sulfur compounds, such as inorganic sulfate, repressed enzyme synthesis while others, such as methionine, did not. Tyramine caused derepression of enzyme synthesis, which is repressed by inorganic sulfate. The present work showed that, although tyramine readily derepressed arylsulfatase synthesis, metabolites of tyramine in either the wild-type or mutant strains did not, so that the derepression is due to the particular structure of tyramine. Kinetic studies on the cells indicated that incorporation of sulfur into protein and enzyme synthesis occurred on supply of either a sulfur compound, which did not cause repression, or of tyramine, which caused derepression, irrespective of the type of sulfur compound added, if any.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.