Isolation and expression of the Bradyrhizobium japonicum adenylate cyclase gene (cya) in Escherichia coli

A 5.0-kilobase-pair HindIII fragment of Bradyrhizobium japonicum DNA containing the cya gene which encodes adenylate cyclase was isolated as an insert in pBR322, using marker rescue of the maltose-negative phenotype of an Escherichia coli cya mutant for identification. The isolated B. japonicum DNA fragment was capable of reversing the pleiotropic phenotype of cya mutations when inserted in either orientation in the HindIII site of pBR322. The complemented E. coli strains produced high levels of cyclic AMP. No sequence homology between the B. japonicum cya gene and that of E. coli was detected by hybridization analysis.     

Effect of mutations for adenylate cyclase (cya) and the cyclic adenosine monophosphate receptor protein (cgp) on the gene expression of nucleoside catabolism in Escherichia coli]

The effect of cya and crp mutations on the expression of the activity of nucleoside catabolizing genes has been studied in Escherichia coli. It is found that cya and crp mutants lose their ability to grow on nucleosides as carbon sources in spite of the preservation of the basal levels of nucleoside catabolizing enzymes, found in cell-free extracts of cya and crp mutants. It is shown that cya and crp mutations completely release the influence of the regulatory gene cytR on the activity of uridine phosphorylase (udp gene) and thymidine phosphorylase (tpp gene). On this ground it is assumed that the cytR gene product acts at the level of promotors of the corresponding structural genes, causing their insensitivity to the positive action of cAMP--CRP complex. The same data concerning the effect of cya and crp mutations on cytR regulation have been reported [8], but these authors favoured the hypothesis that the cytR gene product is a repressor protein, which binds to the specific operator.     

Escherichia coli cells with mutations in the gene for adenylate cyclase (cya) exhibit a heat shock response

Abstract Adenylate cyclase mutants of Escherichia coli showed the heat-shock response. The heat-shock response was studied in two different mutants and in different growth media, including rich and minimal media. These results are in disagreement with the proposal that the cya gene regulates the expression of the heat-shock genes.     

Control mechanism of the Escherichia coli K-12 cell cycle is triggered by the cyclic AMP-cyclic AMP receptor protein complex.

The role of cyclic AMP (cAMP) in the cell cycle of Escherichia coli K-12 was studied in three mutant strains. One was KI1812, in which the cya promoter is replaced by the lacUV5 promoter. In KI1812, isopropyl-beta-D-thiogalactopyranoside induced the synthesis of cya mRNA, and at the same time cell division was inhibited and short filaments containing multiple nuclei were formed. The other strains were constructed as double mutants (NC6707 cya sulB [ftsZ(Ts)] and TR3318 crp sulB [ftsZ(Ts)]). In both double mutants, filamentation was repressed at 42 degrees C, but it was induced again by addition of cAMP in strain NC6707 and introduction of pHA7 containing wild-type crp in TR3318. These results indicate that lateral wall synthesis in the E. coli cell cycle is triggered by the cAMP-cAMP receptor protein complex.     

Cell filamentation in an Escherichia coli K-12 fic mutant caused by theophylline or an adenylate cyclase gene (cya)-containing plasmid

Abstract The bioconversion of 17α-ethynyl steroids was effected with 11α-hydroxylase of Rhizopus nigricans . 7β-Hydroxyethisterone was found after the bioconversion of ethisterone, and 10β- and 6β-hydroxy derivatives after the bioconversion of norethisterone. It seems that the ethynyl group prevents steroid-enzyme binding in the normal mode and thus inhibits the formation of an 11α-hydroxylated product.     

A monoclonal antibody that inhibits cyclic AMP binding by the Escherichia coli cyclic AMP receptor protein

The monoclonal antibody (mAb) 64D1 was found to inhibit cAMP binding by the cAMP receptor protein (CRP) from Escherichia coli (Li, X.-M., and Krakow, J. S. (1985) J. Biol. Chem. 260, 4378-4383). CRP is relatively resistant to attack by the Staphylococcus aureus V8 protease, chymotrypsin, trypsin, and subtilisin whereas both mAb 64D1-CRP and cAMP-CRP are attacked by these proteases yielding N-terminal core fragments. The fragment patterns resulting from proteolysis of mAb 64D1-CRP and cAMP-CRP differ indicating that the CRP in each complex is in a different conformation. The data presented indicate that the preferred conformation of the antigenic site for mAb 64D1 is present in unliganded CRP. Binding of mAb 64D1 to CRP is inhibited at high cAMP concentration. Formation of a stable cAMP-CRP-lac P+-RNA polymerase open promoter complex resistant to dissociation by mAb 64D1 occurs at a much lower cAMP concentration. The observed increase in resistance to mAb 64D1 may reflect a possible conformational change in CRP effected by contact with RNA polymerase in the open promoter complex.     

Differential regulation by cyclic AMP of starvation protein synthesis in Escherichia coli

Of the 30 carbon starvation proteins whose induction has been previously shown to be important for starvation survival of Escherichia coli, two-thirds were not induced in cya or crp deletion mutants of E. coli at the onset of carbon starvation. The rest were induced, although not necessarily with the same temporal pattern as exhibited in the wild type. The starvation proteins that were homologous to previously identified heat shock proteins belonged to the latter class and were hyperinduced in delta cya or delta crp mutants during starvation. Most of the cyclic AMP-dependent proteins were synthesized in the delta cya mutant if exogenous cyclic AMP was added at the onset of starvation. Furthermore, beta-galactosidase induction of several carbon starvation response gene fusions occurred only in a cya+ genetic background. Thus, two-thirds of the carbon starvation proteins of E. coli require cyclic AMP and its receptor protein for induction; the rest do not. The former class evidently has no role in starvation survival, since delta cya or delta crp mutants of either E. coli or Salmonella typhimurium survived starvation as well as their wild-type parents did. The latter class, therefore, is likely to have a direct role in starvation survival. This possibility is strengthened by the finding that nearly all of the cya- and crp-independent proteins were also induced during nitrogen starvation and, as shown previously, during phosphate starvation. Proteins whose synthesis is independent of cya- and crp control are referred to as Pex (postexponential).