A bacterium, strain PC-07, previously isolated as part of a coculture capable of growing on p-cresol under anaerobic conditions with nitrate as the acceptor was identified as an Achromobacter sp. The first enzyme of the pathway, p-cresol methylhydroxylase, which converts its substrate into p-hydroxybenzyl alcohol, was purified.The enzyme had an Mr of 130,000 and the spectrum of a flavocytochrome. It was composed of flavoprotein subunits of Mr 54,000 and cytochrome subunits of Mr 12,500. The midpoint redox potential of the cytochrome was 232 mV. The-Km and kcat for p-cresol were 21 ,uM and 112 s-1 respectively, and the Km for phenazine methosulfate, the artificial acceptor used ig the assays, was determined to be 1.7 mM. These properties place the enzyme in the same class as the p-cresol methylhydroxylases from aerobically isolated Pseudomonas spp.
The survival under starvation conditions of two selected strains of marine bacteria, a yellow Pseudomonas sp. (strain 95A) and an unidentified oxidative peritrichate Gram negative rod (strain 41), was investigated. The 50% survival times of suspensions in phosphate buffer depended on cell density and were often more than 20 d. A capacity to scavenge atmospheric nitrogenous compounds led to a marked increase in the viability of cell suspensions of 104 cells/ml. Intracellular poly‐β‐hydroxybutyrate (PHB) prolonged the survival of strain 95A. Strain 41 contained more intracellular protein and this was degraded during starvation in ammonia‐free air. Prolonged survival was not explicable in terms of low adenylate charge states. The ‘maintenance energy’requirements of strains 95A and 41 in chemostat cultures were 0.042 and 0.04 g glucose/g dry wt/h respectively, compared with dilution‐rate‐dependent values of 0.051 to 0.856 for Escherichia coli. The low maintenance energy requirements would not alone explain the long viability. Thus no peculiar physiological property such as nitrogen‐scavenging, ability to survive at the expense of intracellular PHB or protein, abnormally low cellular protein content, low maintenance energy requirements or a low adenylate charge state fully account for the starvation resistance of these marine bacteria.
A bacterium capable of growth on 4-hydroxyacetophenone was isolated from soil and identified as an Alcaligenes sp. Intact cells rapidly oxidized (4-hydroxybenzoyl)methanol, 4-hydroxybenzoate and protocatechuate as well as the growth substrate, and also converted the substrate analogue (4-methoxybenzoyl)methanol to 4-methoxybenzoic acid. When provided with NADH, cell-free extracts oxidized 4-hydroxyacetophenone to 4-hydroxybenzoate and formate, the same products as were formed from (4-hydroxybenzoyl)methanol without NADH. The oxidation of 4-hydroxybenzoate by cell-free extracts required NADPH and the product from both this and protocatechuate oxidation was 3-oxoadipate. A pathway for the catabolism of 4hydroxyacetophenone, by hydroxylation to (4-hydroxybenzoyl)methanol followed by oxidative cleavage to 4-hydroxybenzoate and formate and hydroxylation of the 4-hydroxybenzoate to protocatechuate, is proposed. Oxidation of protocatechuate was by the ortho pathway. The key enzymes in the proposed pathway were induced by growth on 4-hydroxyacetophenone.
Studies were made of the variations in the numbers of viable bacteria in Cardigan Bay seawater as estimated using the spread‐plate technique and various marine agar media, the results being examined using analysis of variance. Use of duplicated 500 ml sample bottles enabled the detection of statistically significant variations between samples both in terms of numbers of viable bacteria present and their different responses to different counting media, but an important micro‐distribution was also indicated. Further studies showed large differences between 25 ml samples taken 10 cm apart: these could not be explained in terms of cell aggregation as presently understood.
The bactericidal activity of Tinopal AN [1,1-bis(3,N-5-dimethyl-benzoxazol-2-yl)-methine p-toluene sulphonate] was shown to be due to a mechanism entirely independent of its inhibitory effects upon NADH dehydrogenase which were reported previously. Whereas the compound had no significant effect upon DNA synthesis in Escherichia coli D22, RNA and protein synthesis were immediately and markedly inhibited. In confirmation, Tinopal AN caused an immediate cessation in inducible beta-galactosidase synthesis in the same organism. An in vitro assay of the transcription of calf-thymus DNA by purified E. coli RNA polymerase showed that this process was inhibited by Tinopal AN.
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