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The Synthesis of Pteroylglutamates in Germinating Pea Cotyledons

其他書名	(microfilm).
出版	National Library of Canada, 1971
URL	http://books.google.com.hk/books?id=KjPttAEACAAJ&hl=&source=gbs_api

註釋The synthesis of pteroylglutamate in pea seeds has been examined over a seven day germination period. It was observed that a rapid synthesis occurred 40-90 hours after the seeds started to imbibe water. The constituents of the pteroylglutamate pool present in the cotyledons were subjected to DEAE-cellulose column chromatography. Identification of individual derivatives was based upon differential microbiological response and co-chromatography with authentic derivatives before and after treatment with y-glutamyl carboxypeptidases from various sources. The three assay bacteria employed in this study were Lactobacillus casei (ATCC 7469), Streptococcus faecalis (ATCC 8043) and Pediococcus cerevisiae (ATCC 8081). The data show that the major constituents of the pteroylglutamate pool were 10-formyltetrahydropteroylglutamate, 5-formyltetra- hydropteroylglutamate and 5-methyltetrahydropteroylglutamate, together with smaller quantities of their glutamyl conjugated forms. Synthesis of these derivatives was inhibited by use of pteroylglutamic acid antagonists such as aminopterin and amethopterin. These compounds also drastically inhibited germination of the seeds and resulted in the appearance of pteroylglutamic acid which was not detected in the absence of these antagonists. Feeding experiments employing pteroylglutamic acid-2-C[superscript 14] and 5-[Methyl-C[superscript 14] tetrahydropteroylglutamic acid indicated that these derivatives were readily utilized in the synthesis of other pteroylglutamates during germination. In contrast, the only pteroylglutamate containing C[superscript 14] following administration of formate-C[superscript 14] was 10-formyltetrahydropteroylglutamate. Despite the rapid synthesis of pteroylglutamates during germination, the levels of 5,10-methylenetetrahydrofolate dehydrogenase remained relatively uniform during the first 7 days of germination. This enzyme, in common with the corresponding dehydrogenases from other sources, was shown to have a requirement for NADP, formaldehyde and tetrahydropteroy lglutamic acid and was stable only when isolated in the presence of ethylenediamine tetraacetate and 2-mercapto- ethanol. The pteroylglutamate product of the reaction was chromatographed on DEAE-cellulose and found to be 10-formyl- tetrahydropteroylglutamate. In experiments with ammonium sulphate fractionated extracts, ability to synthesize the principal endogenous derivatives was determined in reaction systems in which formaldehyde, formate and serine served as 1-carbon donors. In all cases, formyltetrahydropteroylglutamic acid derivatives were formed. This synthesis displayed requirements for the 1-carbon donors. In addition, 5-methyltetrahydropteroy1- glutamic acid was also produced in these experiments. This synthesis required the presence of a reduced pyridine nucleotide but did not display a requirement for the 1-carbon donor. The results are interpreted as indicating that 5,10-methyltetrahydropteroylglutamic acid, forraed from the donors, was oxidized and reduced in these reaction systems. The significance of these derivatives in the 1-carbon metabolism of these germinating tissues is discussed.