Date of Award:

5-1-1983

Document Type:

Dissertation

Degree Name:

Doctor of Philosophy (PhD)

Department:

Biology

Department name when degree awarded

Life Sciences:Biology

Committee Chair(s)

Jon Y. Takemoto

Committee

Jon Y. Takemoto

Committee

T. M. Farley

Committee

G. W. Miller

Committee

S. G. Oberg

Committee

F. J. Post

Abstract

Aspects of glycolate metabolism in the photosynthetic bacterium Rhodopseudomonas sphaeroides were studied to determine the occurrence of the plant-type glycolate pathway in this bacterium. R. sphaeroides, when grown with tartrate or acetate, excreted glycolic acid extracellularly during chemoheterotrophic or photoheterotrophic growth. The amount of glycolic acid excretion increased when cells were incubated with 10 mM α-hydroxy-2-pyridine methane sulphonate (HPMS), an inhibitor of glycolate oxidase. The rates of bacteriochlorophyll a and carotenoid synthesis were inhibited by HPMS; however, addition of glyoxylate, glycine or serine did not restore the original rates of these pigments. Ribulose 1,5-diphosphate carboxylase, ribulose 1,5-disphosphate oxygenase, phosphoglycolate phosphatase, glycolate oxidase, L- alanine:glyoxylate aminotransferase, glycine decarboxylase and serine hydroxymethyl transferase activities required for the formation and conversion of glycolate to serine, via glyoxylate and glycine, were present in the cell-free crude extracts of R. sphaeroides. Crude extracts of R. sphaeroides decarboxylated exogenously supplied [1-14C] glycolate and [1-14C] glycine. HPMS, 2,3-epoxypropionate (glycidate), isonicotinyl hydrazide (INH) and KCN inhibited this decarboxylation, in a manner, indicating that decarboxylation of glycine is the only major site of CO2 release during the glycolate pathway in R. sphaeroides. Short term incorporation of [1,2-14C] glycolate by the whole cells of R. sphaeroides growing aerobically with tartrate resulted in the sequential incorporation of 14C into glyoxylate, glycine and serine. Incorporation of 14C into serine was much less in the cells growing with malate or anaerobically with tartrate and showed correlation with glycine decarboxylase activities. HPMS, glycidate, INH and KCN inhibited incorporation of 14C into different intermediates of the glycolate pathway. HPMS and INH inhibited net 14CO2 fixation by the cells growing aerobically or anaerobically with tartrate or malate. Also, addition of HPMS (2 mM) to the growth medium inhibited the growth rate of aerobic cells growing on malate or tartrate but growth rates of anaerobic cells remained similar. Among the enzyme activities assayed, glycolate oxidase activities were much less in glycine and serine auxotrophs of R. sphaeroides compared to wild type cells, showing the importance of this pathway in supplying glycine and serine in this bacterium.

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