Cytochromes and dehydrogenases in membranes of a new human periodontal bacterial pathogen, Capnocytophaga ochracea.
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Cell Membrane↗
Biomedical subjects
R Calmes
Publications and source records attributed to R Calmes.
Regulation of lactose catabolism in Streptococcus mutans: purification and regulatory properties of phospho-beta-galactosidase.
Phospho-beta-galactosidase (P-beta-gal), the enzyme which catalyzes the first step in the metabolism of intracellular lactose phosphate, occurred at high specific activity in the cytoplasm in 12 of 13 strains of streptococcus mutans grown on lactose but not other carbon sources. The P-beta-gal from S. mutans SL1 was purified 13-fold using diethylaminoethyl-cellulose ion exchange and agarose A--0.5 M molecular exclusion column chromatography. The molecualr weight of the enzyme was estimated to be 40,000, and its pH optimum was 6.5 in three different buffer systems. P-beta-gal activity was inhibited by Co2+, Zn2+, and Cu2+, but other cations, ethylenediaminetetraacetic acid, orthophosphate, and fluoride had no effect upon enzyme activity. The kinetic response of P-beta-gal to a model substrate, o-nitrophenyl-beta-D-galactopyranoside-6-phosphate, obeyed Michaelis-Menten kinetics, and the Km for this substrate was 0.19 mM. In addition to being under genetic control, P-beta-gal activity was regulated by a number of biologically active metabolites. Enzyme activity was inhibited in a sigmoidal fashion by phosphoenolpyruvate. The M 0.5 V value for phosphoenolpyruvate was 2.8 mM, and the Hill coefficient (n) was 3. In addition, P-beta-gal exhibited strong inhibition by ATP, galactose-6-phosphate, and glucose-6-phosphate. In contrast to inhibition of P-beta-gal activity by phosphoenolpyruvate, the inhibition exerted by ATP, galactose-6-phosphate, and glucose-6-phosphate obeyed classical Michaelis-Menten kinetics; the Ki values for these inhibitors were 0.55, 1.6, and 4.0 mM, respectively.
Adenosine Triphosphate↗
Involvement of phosphoenolpyruvate in the catabolism of caries-conducive disaccharides by Streptococcus mutans: lactose transport.
The mechanisms for transport and hydrolysis of lactose were investigated in five cariogenic strains (HS6, AHT, FA1, NCTC 10449, and SL1) representing the four serogenetic groups of Streptococcus mutans. The systems for transport and hydrolysis of lactose had the characteristics of a phosphoenolpyruvate (PEP)-dependent lactose (Lac) phosphotransferase (PT) system and phospho-beta-galactosidase (P-beta-gal), respectively, in all strains tested, except strain HS6. Decryptified cells required PEP and Mg(2+) for transport of the non-metabolizable model beta-galactosides o-nitrophenyl-beta-d-galactopyranoside (ONPG) and thiomethyl-beta-d-galactopyranoside (TMG). Substitution of 2-phosphoglycerate (2-PG) for PEP also stimulated the Lac PT system. Other potential high-energy phosphate donors (adenosine tri-, di-, and monophosphates and guanosine triphosphate) did not stimulate the Lac PT system. Sodium fluoride had no effect upon the PEP-dependent Lac PT system in decryptified cells with PEP as the energy source; however, when 2-PG was used as the energy source, F(-) inhibited ONPG phosphorylation. With intact cells which must generate PEP endogenously, the presence of F(-) in concentration >/= 10 mM completely inhibited the Lac PT system, presumably through inhibition of 2-PG hydrolyase (EC 4.2.1.11; enolase). Both intact and decryptified cells accumulated a phosphorylated derivative of TMG that behaved chromatographically as TMG-phosphate. After alkaline phosphatase treatment, the derivative had an R(f) identical to that of TMG. No beta-galactosidase (beta-gal) activity was detected with ONPG as the substrate; hydrolysis occurred only when ONPG-6-phosphate was supplied as the substrate. Strain HS6 apparently transported lactose by an active transport-type system in which the accumulated intracellular product was the free disaccharide based on the following criteria: (i) ONPG transport and hydrolysis in decryptified cells was not stimulated by PEP; (ii) ONPG hydrolysis occurred in the absence of PEP; and (iii) ONPG-6-phosphate was not hydrolyzed. These data indicate that, in all strains tested except strain HS6, lactose transport was mediated by a PEP-dependent Lac PT system, resulting in accumulation of lactose-phosphate that was hydrolyzed by an enzyme similar to the P-beta-gal of group N streptococci and Staphylococcus aureus; conversely, strain HS6 transported and hydrolyzed lactose by a PEP-independent transport system and beta-gal, respectively.
Biological Transport↗
Fatty acid transport by the lipophilic bacterium Nocardia asteroides.
Hexadecanoate was translocated in Nocardia asteroides by a constitutive transport system(s), which transported short, medium, and long-chain fatty acids. Inhibition of hexadenocanoate transport by homologues suggested that at least two systems are present: one specific for short-chain fatty acids and the other specific for medium- and long-chain fatty acids. Saturation kinetics typical of a carrier-mediated transport system (Kt = 870 muM)were observed, and concentration of fatty acids against a gradient was achieved. Inhibitor studies indicated that free sulfhydryl groups, a functional respiratory chain, and energy are required for translocation. Efflux of [14C]hexadecanoate in the presence of excess unlabeled hexadecanoate or 2,4-dinitrophenol and the cytoplasmic localization of acyl-coenzyme A synthetase (acid:coenzyme A ligase [adenosine monophosphate]; EC 6.2.1.3) (Calmes and Deal, 1973) are consistent with the hypothesis that fatty acids are transported and released intracellularly as free fatty acids.
Adsorption↗