PubMed Health⌕ Search

Biomedical subjects

N Guiso

Publications and source records attributed to N Guiso.

84 records · Page 5Linked to original sources
PubMed1982

Multiple regulation of the activity of adenylate cyclase in Escherichia coli.

We have studied the correlation between the activities of adenylate cyclase (ATP pyrophosphatelyase-(cyclizing); EC 4.6.1.1) and in vivo rates of synthesis and intracellular concentrations of adenosine 3',5' cyclic monophosphate (cAMP) under various growth conditions in wild-type Escherichia coli and in mutants lacking or overproducing the cAMP receptor protein (CAP). We showed that when wild-type bacteria are grown in the presence of a variety of carbon sources the intracellular concentrations of cAMP are inversely related to the adenylate cyclase activities determined in permeabilized cells, suggesting that the carbon source-dependent modulation of cAMP levels is not directly related to the regulation of adenylate cyclase activity. In mutants lacking functional CAP (crp) the in vivo rates of cAMP synthesis are several hundred-fold higher than in the wild-type parent without a parallel increase of adenylate cyclase activities. In a strain carrying multiple copies of the crp gene and overproducing CAP the activity of adenylate cyclase is severely inhibited, although the in vivo rate of cAMP synthesis is similar to the parental strain. We interpret these results as indicating that CAP controls mainly the activity rather than the synthesis of adenylate cyclase.

Adenylyl Cyclases↗
PubMed1982

crpX mutants of Escherichia coli K12: specific regulatory effects of altered cyclic AMP receptor proteins.

We attempted to correlate structural modifications of the adenosine 3',5' cyclic monophosphate (cAMP) receptor protein (CAP), to changes in some of its in vivo regulatory functions such as (i) stimulation of the lactose operon expression and (ii) control of adenylate cyclase activity. A radioimmunological procedure was used to study the structure of CAP synthesized by three mutants (crpX) grown under various conditions, in the presence or absence of endogenous or exogenous cAMP. In one mutant CAP appears to be sensitive to thermal inactivation. In another mutant CAP is particularly sensitive to degradation in the absence of cAMP; this degradation is enhanced by high temperature and during stationary phase of growth, and prevented by the addition of glucose. Functional alterations of CAP were not found to follow structural changes strictly. In the crpX mutants and in strains carrying the crp+ or other crp allele, the stimulation of the lactose operon expression and the modulation of the in vivo rates of cAMP synthesis appear to vary in parallel, favoring an indirect mechanism of regulation of adenylate cyclase by CAP.

Adenylyl Cyclases↗
PubMed1978

An immunological determination of specific activities of enzymes: its use in quantification of cross reactivity between enzymes of different origins.

An immunological method is presented which enables the determination of the specific activity of a pure enzyme without its extensive purification. The method consists essentially in the specific fixation of immunologically related enzymes to an immunoadsorbent containing specific antibodies raised against the wild-type form of the enzyme. We applied this method to determine the specific activity of plasmid-coded beta-galactosidases and to quantify the extent of cross-reaction between these enzymes and Escherichia coli beta-galactosidase.

Animals↗
PubMed1976

Expression and regulation of lactose genes carried by plasmids.

A number of plasmids carrying the lactose character have been studied. All of the plasmids examined so far code for proteins essential for lactose utilization, i.e., beta-galactosidase and galactoside permease. None of them carries enzymatically or immunologically detectable thiogalactoside transacetylase. The expression of the two enzymes is both negatively and positively controlled: they are inducible by different galactosides and are sensitive to catabolite repression. Since the plasmid-coded lactose systems have many features in common with the Escherichia coli lactose operon, it is suggested that the plasmids could have acquired the lactose genes from an E. coli chromosome.

Acetyltransferases↗
PubMed1975

The threonine-sensitive homoserine dehydrogenase and aspartokinase activities of Escherichia coli K12. Distribution and accessibility to antibodies of some epitopes of the bifunctional enzyme.

In the presence of l-threonine, the allosteric effector, most of the antigenic determinants situated in the aspartokinase region of the wild-type enzyme become unavailable to the antibodies raised against a fragment of the enzyme containing this region and devoid of homoserine dehydrogenase activity. The cross-reactivities of the antibodies raised against this fragment (extracted from a nonsense mutant) and a fragment endowed with homoserine dehydrogenase activity but devoid of aspartokinase activity (obtained by limited proteolysis) with the corresponding antigens were studied. The conclusion is drawn that the two fragments, which share an overlapping sequence of molecular weight about 17,000, share at least two antigenic determinants.

Alcohol Oxidoreductases↗
PubMed1975

Evolution of biosynthetic pathways: immunological approach.

Through the use of specific immunoadsorbent columns, it is shown that Escherichia coli aspartokinase I-homoserine dehydrogenase I, aspartokinase II-homoserine dehydrogenase II, aspartokinase III, and homoserine kinase, enzymes involved in the same complex biosynthetic pathway, share antigenic determinants. This raises the question of a common origin for the four cibtenoirart kinases. (Aspartate kinase or ATP:L aspartate 4-phosphotransferase, EC 2.7.2.4; homoserine dehydrogenase or Lhomoserine:NADP oxidoreductase, EC 1.1.1.3; homoserine kinase or ATP:L-homoserine O-phosphotransferase, EC 2.7.1.39.)

Alcohol Oxidoreductases↗
PubMedDate not supplied

Cloning and expression in Escherichia coli of the TL-DNA gene 4 of Agrobacterium tumefaciens under the control of the PR promoter of bacteriophage lambda.

A plasmid was constructed that directs expression of the TL-DNA gene 4 protein in E. coli. The different steps of the construction were as follows: i) a region of gene 4 encoding the amino-terminal portion of the protein was fused in frame to DNA encoding an enzymatically active carboxy-terminal fragment of beta-galactosidase. The hybrid gene was poorly expressed from the upstream lambda PL promoter carried by the vector. ii) in order to generate an efficient procaryotic ribosome binding site, a DNA fragment carrying the lambda PR promoter with the nearby Shine-Dalgarno (SD) sequence of gene cro was placed in front of the gene 4-lacZ fusion. A recombinant plasmid, termed pGV793, that expressed efficiently a fused protein 4-beta-galactosidase was identified among the Lac+ clones. DNA sequencing analysis showed that pGV793 carried a hybrid ribosome binding site composed of the cro SD sequence, a five bp sequence and the ATG codon of gene 4. Plasmid pGV793 directed the synthesis of three polypeptides of molecular weight 132 Kd, 126 Kd and 122 Kd that carried beta-galactosidase antigenic determinants. The largest polypeptide had the expected size for the hybrid protein. The fusion proteins which accounted for about 0.5% of the total cellular proteins were purified by immunoadsorption using anti-beta-galactosidase antiserum. iii) the complete gene 4 coding sequence was reconstituted, with the lambda PR promoter in place. The resulting pGV822 plasmid expressed a polypeptide whose molecular weight 27 Kd corresponded to the expected size for the gene 4 product. The pI was about 7.

Bacteriophage lambda↗