A species of Rhodopseudomonas that grows under strict anaerobic conditions in the dark and requires CO was isolated from lake and pond sediments. Although anaerobic growth in the dark occurs in a chemically defined mineral medium with CO as the only carbon and energy source, growth is stimulated by adding trypticase. Under these conditions, cells exhibit a generation time of 6.7 hr and reach a final concentration of I to 3 X 109 cells per ml of liquid medium. Resting suspensions of CO-grown cells metabolize about 6.7 Vmol of CO per mg of protein in 1 hr and produce equimolar amounts of CO2 and H2 according to the equation CO + H210 -CO2 + H2. As predicted by this equation, when cells were suspended in tritium-labeled water containing potassium phosp ate buffer at pH 7.0 and incubated with pure CO, 3H2 gas was produced at linear rate with a constant specific activity. Carbon monoxide is a natural by-product in a variety of biologic and chemical reactions that occur in nature (1-5). Once formed, the gas may be oxidized to carbon dioxide by photochemical reactions in the troposphere (6). Evidence that biologic reactions also occur for oxidation and/or utilization of CO has been more difficult to obtain (7,8). Both plant and bacterial systems have been implicated.The ability of leaf tissue from young bean plants to oxidize CO to CO2 in light led to speculation that plants play an important role in removing CO from the atmosphere. Nevertheless, since the reaction was not observed when other tissfies were used (2), the ability to oxidize CO may not be a general property of green plants. In addition, it has been reported that a variety of aerobic and anaerobic bacteria metabolize carbon monoxide. However, in these studies performed with suspensions of whole cejls (9) or protein extracts (10-13), the reaction(s) occurred slowly and amounts of CO oxidized to CO9 were small. Likewise a few studies (14,15) suggested certain aerobic microorganisms could grow with CO. Unfortunately, important experimental details were lacking and it was not possible to determine if the cells grew methylotrophically (7, 8), i.e., with CO as a source of carbon and energy, or if the gas was cometabolized (10, 11) with other substrates required for cell growth. It has been generally accepted that two species of methanogenic bacteria can oxidize CO to CO2 under anaerobic conditions, and with H2 can form methane gas and grow (16)(17)(18) monoxide-utilizing microorganisms were selectively isolated from anaerobic enrichment cultures. About 50 ml of reduced liquid medium (described below) was prepared, but without adding trypticase, and inoculated with a 1 g sample of anaerobic, black sediment from Crystal Lake, Urbana, Ill. or from a duck pond located outside Champaign, Ill. Enrichment cultures were incubated at 340 under a stream of sterile, oxygenfree H2-CO gas (80:20 mixture). When turbid growth developed, a sample of cells was obtained from the enrichment culture by means of a platinum inoculation loop and transferred into 50 ml of ster...
Purple nonsulfur photosynthetic bacteria were cultured anaerobically in the absence of light by a modification of the Hungate technique. Growth was slow and resembled that of fastidious anaerobes; on yeast extract-peptone-agar medium, each cell produced about 16 descendants in 15 to 20 days. Growth was stimulated by addition of ethyl alcohol, acetate and H2, or pyruvate and H2. Cells grown in the presence of pyruvate and H2 produced acetate and C02; each cell produced approximately 10 descendants in 24 hr under anaerobic, dark conditions. Spectrophotometric evidence obtained from cells which were the product of five generations suggests no difference between the bacteriochlorophyll and carotenoids synthesized by cells grown anaerobically under dark or light conditions. Likewise, the ultrastructure of the photosynthetic apparatus in cells grown anaerobically in the dark and in the light appears similar.
The pathway of pulcherriminic acid synthesis in Bacillus subtilis strains AM and AM-Li1 (a leucine-requiring auxotroph) was investigated. Determinations of radioactivity in pulcherriminic acid synthesized by cells growing in media containing "C-labeled amino acids indicated that B. subtilis produced pulcherriminic acid from L-leucine. The organism utilized the carbon skeletons of two L-leucine molecules to synthesize one molecule of pulcherriminic acid. Similar results were obtained with starved cell suspensions. Growing cells formed significant amounts of pulcherriminic acid only in media including a carbohydrate such as starch. However, carbohydrate carbon was not required for the synthesis of pulcherriminic acid molecules. Data obtained with cell suspensions supported the hypothesis that cyclo-L-leucyl-L-leucyl is an intermediate in pulcherriminic acid biosynthesis and indicated that molecular oxygen is required for the conversion of cyclo-L-leucyl-L-leucyl to pulcherriminic acid. A pathway for the synthesis of pulcherrimin from L-leucine in B. subtilis is proposed.
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