Molecular cloning and expression of a cDNA sequence encoding histidinol phosphate aminotransferase from Nicotiana tabacum

A Nicotiana tabacum cDNA sequence encoding histidinol phosphate aminotransferase (HPA) was isolated by functional complementation of an Escherichia coli histidine auxotroph (UTH780). The enzymatic assay has confirmed that the isolated cDNA encodes a functional HPA protein. Amino acid sequence alignm...

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Bibliographic Details
Published in:Plant molecular biology 1998-08, Vol.37 (6), p.1013-1022
Main Authors: El-Malki, F. (Vrije Univ. Brussel, Sint-Genesius Rode (Belgium). Lab. for Plant Genetics), Frankard, V, Jacobs, M
Format: Article
Language:English
Subjects:
ADN
Amino Acid Sequence
Amino acids
AMINOTRANSFERASAS
AMINOTRANSFERASE
AMINOTRANSFERASES
Base Sequence
CLONACION MOLECULAR
CLONAGE MOLECULAIRE
Cloning
Cloning, Molecular
DNA
DNA, Complementary
E coli
Enzymes
Escherichia coli - genetics
EXPRESION GENICA
EXPRESSION DES GENES
GENE EXPRESSION
Gene Library
Genes, Plant
Genetic Complementation Test
Histidine - biosynthesis
MOLECULAR CLONING
Molecular Sequence Data
Nicotiana - enzymology
Nicotiana - genetics
NICOTIANA TABACUM
NUCLEOTIDE SEQUENCE
Plants, Toxic
Recombinant Proteins - biosynthesis
RNA, Messenger - genetics
RNA, Plant - genetics
SECUENCIA NUCLEOTIDICA
Sequence Analysis, DNA
Sequence Homology, Amino Acid
SEQUENCE NUCLEOTIDIQUE
Transaminases - biosynthesis
Transaminases - genetics
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Description
Summary:A Nicotiana tabacum cDNA sequence encoding histidinol phosphate aminotransferase (HPA) was isolated by functional complementation of an Escherichia coli histidine auxotroph (UTH780). The enzymatic assay has confirmed that the isolated cDNA encodes a functional HPA protein. Amino acid sequence alignment of the HPA protein from N. tabacum, Saccharomyces cerevisiae and E. coli revealed that, despite the low degree of identity, some residues were found to be highly conserved. The predicted protein contains a transit peptide sequence at the amino-terminal end, suggesting a chloroplastic localization of the HPA enzyme. Western blot analysis demonstrated that the deduced HPA protein and the mature HPA protein have an apparent molecular mass of about 45 kDa and 40 kDa respectively. Gene copy number estimation by Southern analysis indicates the presence of at least two genes per haploid genome coding for this protein in Nicotiana sp. From northern analysis results, the gene seems to be highly expressed in green tissues and the detected transcript showed a single band of expected molecular size.
ISSN:0167-4412
1573-5028
DOI:10.1023/A:1006007125448