ISSN:
1573-5028
Keywords:
cDNA
;
gene expression
;
hydroxypyruvate reductase
;
light regulation
;
peroxisomal enzyme
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
Notes:
Abstract A full-length cDNA encoding NADH-dependent hydroxypyruvate reductase (HPR), a photorespiratory enzyme localized in leaf peroxisomes, was isolated from a λgt11 cDNA library made by reverse transcription of poly(A)+ RNA from cucumber cotyledons. In vitro transcription and translation of this clone yielded a major polypeptide which was identical in size, 43 kDA, to the product of in vitro translation of cotyledonary poly(A)+ RNA and subsequent immunoprecipitation with HPR antiserum. Escherichia coli cultures transformed with a plasmid construct containing the cDNA insert were induced to express HPR enzyme activity. RNA blot analysis showed that HPR transcript levels rise significantly in the first eight days of light-grown seedling development. This closely resembles the pattern seen for HPR-specific translatable mRNA. DNA blot analysis indicated that a single HPR gene is likely present per haploid genome. Nucleotide sequence analysis revealed an open reading frame of 1146 bases which encodes a polypeptide with a calculated molecular weight of 41.7 kDa. The derived amino acid sequence from this open reading frame is 26% identical and 50% similar to the amino acid sequence of the E. coli enzyme phosphoglycerate dehydrogenase, which catalyzes a similar reaction and functions in a related pathway. Statistical analyses show that this similarity is significant (z〉10). The derived amino acid sequence for HPR also contains the characteristics of an NAD-binding domain.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF00016133
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