The transmissible plasmid pAMbeta1, which codes for erythromycin and lincomycin resistance, was conjugally transferred from a Lancefield group F Streptococcus to a strain of Streptococcus avium. Both organisms served as pAMbeta1 donors for three strains of Lactobacillus casei. Introduction of pAMbeta1 into one of the L. casei strains caused the organism to lose its native 6.7 X 10(6)-dalton plasmid. Loss of the native plasmid produced no alterations in the organism's growth characteristics or fermentation pattern.
Certain strains of Lactobacillus casei can grow at the expense of one or more pentitols. These microorganisms possess a pentitol phosphate pathway that appears to be analogous to the hexitol phosphate pathway found in many facultatively anaerobic bacteria. Pentitol is transported into the cell by a phosphoenolpyruvate phosphotransferase system that converts it to pentitol phosphate, whereupon a specific dehydrogenase oxidizes the intermediate product to protein per ml with 0.01 M potassium phosphate buffer, pH 7.0. Protein contents of such suspensions were determined by the method of La Riviere (6). Cell extracts for enzyme assays were prepared by suspending cell pastes (2-4 g wet weight) in 10 ml of potassium phosphate buffer containing 10 mM 2-mercaptoethanol and treating them with an icewater-cooled Branson model 350 ultrasonic cell disrupter operating at 175 W for 10 min. The crude extract was centrifuged for 20 min at 20,000 X g and the clarified supernatant fluid was decanted and assayed for enzyme activity. Protein content of cell extracts was measured by the biuret technique (7).Enzyme Assays. Pentitol phosphate dehydrogenase activity was measured by following the rate of NADH oxidation at 340 nm in a Gilford model 2400S recording spectrophotometer using xylulose-or ribulose-5-P (Xu5P or Ru5P) as substrate. The reaction mixture contained the following: potassium phosphate buffer, pH 6.5, 50 mM; 2-mercaptoethanol, 5 mM; NADH, 0.1 mM; MgCl2, 50 mM; Xu5P, 2 mM, or Ru5P, 5 mM; sufficient extract to give a rate of 0.2-0.4 absorbance units/min; and water to 1 ml. Endogenous NADH oxidation was measured for 2 min in the absence of substrate to obtain a basal value and the reaction was initiated by the addition of Ru5P or Xu5P. All dehydrogenase values are corrected for NADH oxidase activity.Activity of the phosphoenolpyruvate phosphotransferase system (PTS) was measured by the Maryanski and Wittenberger (8) modification of a previously published procedure (9). Duplicate 10-Al samples were withdrawn at 5-min intervals over a 30-min period from a reaction mixture containing Tris.HCl buffer, pH 7.5, 100 mM; phosphoenolpyruvate (PEP), 10 mM; MgCl2, 5 mM; 2-mercaptoethanol, 1 mM; [U-14C]-xylitol (Amersham/Searle), 5 mM (5 X 105 cpm) or [1-14C]-ribitol (New England Nuclear), 5 mM (4 X 105 cpm); extract, 4-6 mg of protein; and distilled water to 0.5 ml; the incubation temperature was 300. The samples were applied over the surface of DE81 filter paper pads (Whatman, diameter 2.5 cm), quickly dried with a stream of air, and washed twice with 5 ml of distilled water. After complete drying, the amount of radiolabeled pentitol phosphate absorbed to the pad was measured in a Beckman model LS-350 liquid scintillation counter. The remainder of the PTS reaction mixture was incubated an additional 4 hr, after which time the solution was deproteinized by the addition of 100 ,ul of 10% trichloroacetic acid. The solution was clarified by centrifugation, neutralized with 0.5 M NaOH, and stored at -40°until used for thin-l...
Antiserum prepared against a Streptococcus faecalis fructose diphosphate aldolase (EC 4.1.2.13) was used to measure the extent of immunological homology between the reference enzyme and aldolases of various streptococci and gram-positive nonsporeforming anaerobic bacteria. The majority of streptococci surveyed were isolated from the oral cavity or blood samples. Strains within the species Streptococcus mutans Clarke and Streptococcus mitis (mitior) possessed aldolases that exhibited marked antigenic heterogeneity and thereby contrasted sharply with previously studied species of Streptococcus (London and Kline, 1973). Varying degrees of cross-reactivity were observed between the aldolases of certain species of Eubacterium, Butyribacteriurn, and Propionibacterium and the anti-streptococcal aldolase serum. Evidence is also presented confirming an earlier report by Neimark (1974) that aldolases of Acholeplusrna species react with the anti-S. faecalis aldolase serum.In a previous study, the fructose diphosphate (FDP) aldolase (EC 4.1.2.13) of Streptococcus faecalis ATCC 27792 (formerly strain MR) was used as a reference point to establish evolutionary relationships among various homofermentative lactic acid bacteria by determining the degree of immunological reactivity between their aldolases and a specific anti-S. faecalis aldolase serum (18). Despite significant differences in the quaternary arrangement of the subunits of this enzyme (17), their primary structure was sufficiently conserved to permit a comparison of their antigenic determinants. A preliminary phylogenetic map of streptococci, lactobacilli, and pediococci was constructed from both qualitative and quantitative immunological data (18).In the present study, the streptococcal branch of the phylogenetic map is confirmed and extended with new immunological data for aldolases of previously untested strains. Also, immunological heterogeneity has been detected among the aldolases from strains of streptococci within the species Streptococcus mutans and Streptococcus mitis (mitior). The data indicate that neither species is homogeneous. A s_trong reaction between the aldolase of Butyribacterium rettgeri and anti-S. faecalis aldolase serum prompted the survey reported here for other immunologically related aldolases among grampositive nonsporeforming anaerobic bacteria. The survey includes organisms belonging to the genera Eu b ac terium, Propioni bact erium , and Arac hnia. MATERIALS AND METHODSOrganisms. The strain designations of the organisms are listed in Table 1 Maintenance and cultivation of t h e microorganisms. The streptococci were all maintained on N.I.H. thioglycolate broth (Difco Laboratories, Detroit, Mich.). For enzymological and immunological studi'es, 2 liters of complex streptococcal medium (16) was inoculated with 10 ml of a 12-h-old culture of the appropriate organism and incubated at 37 C for 18 h. Cells were harvested by centrifugation and washed twice with cold 0.05 M potassium phosphate buffer, pH 7.0. Washed cell pellets were stored at -...
Strains of Lactobacillus casei capable of growing on either ribitol or xylitol carry out a heterolactic fermentation producing ethanol, acetate, and a mixture of D-and L-lactate. Following conversion of the pentitols to ribulose 5-phosphate or xylulose 5-phosphate via enzymatic steps unique to these organisms, the intermediate products are further metabolized by enzymes of the pentose pathway. The initial enzymes of the pathway, i.e., pentitol:phosphoenolpyruvate phosphotransferase and pentitol phosphate dehydrogenase, do not appear to be stringently regulated by glucose or intermediate products of glycolysis.
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.
customersupport@researchsolutions.com
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.