Irwin, Stephen D. (1989) Molecular cloning of the major castor bean 25 albumin precursor. PhD thesis, University of Warwick.

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Abstract

Rabbit antibodies raised against the glutamine-rich castor bean 2S albumin, a heterodimer of 4kDa and 7kDa polypeptides, had previously been shown to immunoprecipitate a 34kDa polypeptide from the total products formed when castor bean endosperm mRNA was translated in vitro.

A cDNA library was constructed using maturing castor bean endosperm mRNA as template. Clones containing sequences complementary to albumin mRNA were isolated by hybridisation using as a probe a mixture of synthetic oligonucleotides representing sequences predicted for a peptide present In the 2S albumin large subunit. The nucleotide sequence contained an open reading frame encoding a preproprotein of 258 amino acid residues. The preproprotein included both polypeptides of the previously sequenced 2S albumin. In addition, this precursor Included two further glutamine-rich sequences which, in terms of their size and conserved cysteine residues typically found in seed proteins of the 2S albumin superfamily, possibly represent the small and large polypeptide subunits of a second heterodimeric storage protein. A post-translational processing scheme has been proposed which would result in a single preproprotein generating two distinct heterodimeric 2S albumins. Studies on the protein bodies of castor bean endosperm cells suggested the existence of a second 2S albumin.

Genomic Southern blots indicated that at least 4 genes encode Ricinus 2S albumin precursors, and a Northern developmental analysis suggested that expression of such genes is temporally regulated. Genomic clones were isolated that represented the albumin genes and one of these was sequenced fully, in order to study the nature of the promoter control regions further. On comparison with the upstream regulatory regions of other 2S albumin genes, a region of high homology spanning 70 base pairs was found. This included a repetitive domain noted in many seed protein genes and implicated in storage protein promoter specificity. It was also located in ricin found likewise in castor bean endosperm cells.

Item Type:	Thesis or Dissertation (PhD)
Subjects:	Q Science > QH Natural history > QH426 Genetics
Library of Congress Subject Headings (LCSH):	Castor oil plant -- Genetics, Albumins, Castor oil plant -- Seeds -- Storage, Plant genetic regulation, Promoters (Genetics), Molecular cloning
Official Date:	September 1989
Dates:	Date Event September 1989 Submitted
Institution:	University of Warwick
Theses Department:	Department of Biological Sciences
Thesis Type:	PhD
Publication Status:	Unpublished
Supervisor(s)/Advisor:	Lord, Mike (J. Mike)
Sponsors:	Science and Engineering Research Council (Great Britain)
Format of File:	pdf
Extent:	293 leaves : illustrations, charts
Language:	eng

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