Kinetics of L-tryptophan production from indole and L-serine catalyzed by whole cells with tryptophanase activity

SummaryPlatelet heterogeneity has been studied with a technique called functional fractionation which employs gentle centrifugation to yield subpopulations (“reactive” and “less-reactive” platelets) after exposure to small doses of aggregating agent. Aggregation kinetics of the different platelet populations were investigated by quenched-flow aggregometry. The large, “reactive” platelets were more sensitive to ADP (Ka = 1.74 μM) than the smaller “less-reactive” platelets (Ka = 4.08 μM). However, their maximal rate of aggregation (Vmax, % of platelets aggregating per sec) of 23.3 was significantly lower than the “less-reactive” platelets (Vmax = 34.7). The “reactive” platelets had a 2.2 fold higher level of cyclic AMP.Platelet glycoproteins were labeled using the neuraminidase-galactose oxidase – [H3]-NaBH4 technique. When platelets were labeled after reversible aggregation, the “reactive” platelets showed a two-fold decrease in labeling efficiency (versus control platelets). However, examination of whole cells or membrane preparations from reversibly aggregated platelets revealed no significant difference in Coomassie or PAS (Schiff) staining.These results suggest that the large, “reactive” platelets are more sensitive to ADP but are not hyperaggregable in a kinetic sense. Reversible aggregation may cause a re-orientation of membrane glycoproteins that is apparently not characterized by a major loss of glycoprotein material.

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Kinetics of substrate oxidation by whole cells and cell membranes of

FEMS Microbiology Letters ◽

10.1016/0378-1097(95)00130-w ◽

1995 ◽

Vol 129 (1) ◽

pp. 33-38 ◽

Cited By ~ 25

Author(s):

H CHANG ◽

S MARCELLI ◽

A DAVISON ◽

P CHALK ◽

R POOLE ◽

...

Keyword(s):

Cell Membranes ◽

Substrate Oxidation ◽

Whole Cells ◽

Kinetics Of

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Antimicrobial Properties and Mode of Action of the Pyrrothine Holomycin

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.45.2.532-539.2001 ◽

2001 ◽

Vol 45 (2) ◽

pp. 532-539 ◽

Cited By ~ 71

Author(s):

Brunello Oliva ◽

Alexander O'Neill ◽

Jenny M. Wilson ◽

Peter J. O'Hanlon ◽

Ian Chopra

Keyword(s):

Rna Polymerase ◽

Rna Synthesis ◽

Antimicrobial Properties ◽

Stringent Response ◽

Active Species ◽

Chain Elongation ◽

Whole Cells ◽

E Coli ◽

Eukaryotic Microorganisms ◽

Kinetics Of

ABSTRACT Holomycin, a member of the pyrrothine class of antibiotics, displayed broad-spectrum antibacterial activity, inhibiting a variety of gram-positive and gram-negative bacteria, with the exception ofEnterobacter cloacae, Morganella morganii, andPseudomonas aeruginosa. The antibiotic lacked activity against the eukaryotic microorganisms Saccharomyces cerevisiae and Candida kefyr. Holomycin exhibited a bacteriostatic response against Escherichia coli that was associated with rapid inhibition of RNA synthesis in whole cells. Inhibition of RNA synthesis could have been a secondary consequence of inhibiting tRNA aminoacylation, thereby inducing the stringent response. However, the levels of inhibition of RNA synthesis by holomycin were similar in a stringent and relaxed pair of E. coli strains that were isogenic except for the deletion of therelA gene. This suggests that inhibition of RNA synthesis by holomycin could reflect direct inhibition of DNA-dependent RNA polymerase. Examination of the effects of holomycin on the kinetics of the appearance of β-galactosidase in induced E. colicells was also consistent with inhibition of RNA polymerase at the level of RNA chain elongation. However, holomycin only weakly inhibitedE. coli RNA polymerase in assays using synthetic poly(dA-dT) and plasmid templates. Furthermore, inhibition of RNA polymerase was observed only at holomycin concentrations in excess of those required to inhibit the growth of E. coli. It is possible that holomycin is a prodrug, requiring conversion in the cell to an active species that inhibits RNA polymerase.

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Oxidation-reduction kinetics of cytochrome f in whole cells

Journal of Photochemistry and Photobiology B Biology ◽

10.1016/1011-1344(87)80005-2 ◽

1987 ◽

Vol 1 (1) ◽

pp. 45-60

Author(s):

P. Maróti ◽

D. Mende ◽

W. Wiessner

Keyword(s):

Cytochrome F ◽

Reduction Kinetics ◽

Whole Cells ◽

Oxidation Reduction ◽

Kinetics Of

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Biosynthesis of a mycobacterial lipopolysaccharide. Incorporation of [14C]acyl groups by whole cells in vivo

Biochemical Journal ◽

10.1042/bj1320329 ◽

1973 ◽

Vol 132 (2) ◽

pp. 329-340 ◽

Cited By ~ 10

Author(s):

Koyu Narumi ◽

John M. Keller ◽

Clinton E. Ballou

Keyword(s):

Fatty Acids ◽

Specific Radioactivity ◽

Short Chain Fatty Acids ◽

The Other ◽

Whole Cells ◽

Mycobacterium Phlei ◽

Turn Over ◽

Kinetics Of ◽

Normal Constituent

1. Mycobacterium phlei (A.T.C.C. 356) cells were incubated with 14C-labelled short-chain fatty acids and the 6-O-methylglucose-containing lipopolysaccharides that became esterified with radioactive acyl groups were isolated. The pattern of labelling of these lipopolysaccharides with the different acyl groups, the effects of different conditions on labelling patterns, and the kinetics of the turnover of 14C-labelled acyl groups were studied. 2. The labelling patterns are summarized as follows. [1-14C]Acetate was incorporated into all of the acyl groups. [1-14C]Propionate led to labelling of propionate and succinate, while [1-14C]isobutyrate was incorporated mostly as such, along with a trace amount in iso-octanoate. 3. Under the conditions of the experiments, [1-14C]acetate was rapidly incorporated into succinyl (3-carboxypropionyl) and octanoyl groups, whereas the acetyl groups themselves were labelled more slowly. Radioactivity in propionyl and succinyl groups, originating from [1-14C]propionate, attained maximum values and then gradually decreased in both. Incorporation of [1-14C]isobutyrate proceeded slowly but reached a plateau and remained constant. While n-butyrate is not a normal constituent of methyl-glucose-containing lipopolysaccharides, it was incorporated as such when n-[1-14C]-butyrate was supplied in the medium. 4. [1-14C]Acetyl groups were readily displaced by unlabelled acetate. On the other hand, the specific radioactivity of the succinyl group continued to increase during a 3h incubation with unlabelled succinate. Propionyl and succinyl groups, labelled by [1-14C]propionate, were displaced slowly by unlabelled propionate or succcinate. The isobutyryl group of the lipopolysaccharides did not turn over, in contrast to the results obtained with the other acyl substituents.

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Volume growth, murein synthesis, and murein cross-linkage during the division cycle of Escherichia coli PA3092

Journal of Bacteriology ◽

10.1128/jb.152.3.1248-1254.1982 ◽

1982 ◽

Vol 152 (3) ◽

pp. 1248-1254

Author(s):

A J Olijhoek ◽

S Klencke ◽

E Pas ◽

N Nanninga ◽

U Schwarz

Keyword(s):

Escherichia Coli ◽

Sodium Dodecyl ◽

Volume Growth ◽

Diaminopimelic Acid ◽

Cross Linking ◽

Penicillin Binding Protein ◽

Whole Cells ◽

Cross Linkage ◽

The Cross ◽

Kinetics Of

Cells of Escherichia coli PA3092 were synchronized by centrifugal elutriation. The synchronously growing cells were double labeled with -3H or DL-[meso-2,6-14C]diaminopimelic acid (DAP) at different times. Cells incorporated [3H]DAP at a continuously increasing rate during their cycle, with a maximum occurring at about 30 min before division for trichloroacetic acid-precipitated cells (whole cells) and about 10 min before division for sodium dodecyl sulfate-treated cells (sacculi). This was in good agreement with the observed kinetics of volume growth under these conditions. Furazlocillin, which preferentially interacts with penicillin-binding protein 3, modified the pattern of incorporation of [3H]DAP. Electron microscopy indicated that furazlocillin did not inhibit the initiation of division but rather its completion. In addition, we measured the cross-linking of the murein inserted at different times during synchronous growth. The highest percentages were found to occur around division. At this same time, the cross-linking of old peptidoglycan was found to be decreased.

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Synthesis, transport, and utilization of specific flagellar proteins during flagellar regeneration in Chlamydomonas.

The Journal of Cell Biology ◽

10.1083/jcb.93.3.615 ◽

1982 ◽

Vol 93 (3) ◽

pp. 615-631 ◽

Cited By ~ 41

Author(s):

S P Remillard ◽

G B Witman

Keyword(s):

Gel Electrophoresis ◽

Two Dimensional Gel Electrophoresis ◽

Whole Cells ◽

Complex Protein ◽

Specific Proteins ◽

Dilution Experiments ◽

Flagellar Regeneration ◽

Flagellar Proteins ◽

Polypeptide Chains ◽

Kinetics Of

We labeled gametes of Chlamydomonas with 10-min pulses of 35SO4(-2) before and at various times after deflagellation, and isolated whole cells and flagella immediately after the pulse. The labeled proteins were separated by one- or two-dimensional gel electrophoresis, and the amount of isotope incorporated into specific proteins was determined. Individual proteins were identified with particular structures by correlating missing axonemal polypeptides with ultrastructural defects in paralyzed mutants, or by polypeptide analysis of flagellar fractions. Synthesis of most flagellar proteins appeared to be coordinately induced after flagellar amputation. The rate of synthesis for most quantified proteins increased at least 4- to 10-fold after deflagellation. The kinetics of synthesis of proteins contained together within a structure (e.g., the radial spoke proteins [RSP] ) were frequently similar; however, the kinetics of synthesis of proteins contained in different structures (e.g., RSP vs. alpha- and beta-tubulins) were different. Most newly synthesized flagellar proteins were rapidly transported into the flagellum with kinetics reflecting the rate of growth of the organelle; exceptions included a central tubule complex protein (CT1) and an actinlike component, both of which appeared to be supplied almost entirely from pre-existing, unlabeled pools. Isotope dilution experiments showed that, for most quantified axonemal proteins, a minimum of 35-40% of the polypeptide chains used in assembling a new axoneme was synthesized during regeneration; these proteins appeared to have predeflagellation pools of approximately the same size relative to their stoichiometries in the axoneme. In contrast, CT1 and the actinlike protein had comparatively large pools.

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