The nuclear-coded chloroplast 22-kDa heat-shock protein of Chlamydomonas. Evidence for translocation into the organelle without a processing step.

Abstract

A cDNA clone, pCHS62, was isolated using poly(A)-rich RNA from heat-shocked Chlamydomonas reinhardtii cells. The clone has a length of 1.1 kb and codes for the complete heat-shock protein which was reported to be associated with the grana region of the thylakoid membranes and ascribes protection against photoinhibition during heat-shock. An expression vector prepared in the pUC19 plasmid was used to obtain a fusion protein against which rabbit polyclonal antibodies have been raised. The antibodies react specifically with the heat-shock protein of 22 kDa synthesized in vivo during heat-shock, which is localized in the grana thylakoids, with the in vitro translated product using poly(A)-rich RNA from heat-treated cells as well as with the hybrid release translation product of the pCHS62 clone. The clone was sequenced. It contains a 5' region consisting of 85 nucleotides, an open reading frame of 471 nucleotides and a non-coding 3' region of 600 nucleotides. Northern hybridization indicates a length of 1.7 kb for the messenger RNA of heat-shock protein 22. Analysis of similarity between the derived amino acid sequence of this protein and other heat-shock proteins demonstrates that this protein belongs to the small-molecular-mass plant heat-shock protein family and also shows similarities with animal heat-shock proteins including the presence of a short region possessing similarity with bovine alpha-crystalline as reported for other heat-shock proteins. The molecular mass of the protein as determined from the sequence is 16.8 kDa. Despite its localization in the chloroplast membranes, it does not seem to include a transit peptide sequence, in agreement with previous data. The sequence contains only a short hydrophobic region compatible with its previously reported localization as a thylakoid extrinsic protein.

• Authors: B Grimm, D Ish-Shalom, D Even, H Glaczinski, P Ottersbach, I Ohad, K Kloppstech
• Journal: European journal of biochemistry (Eur J Biochem) Vol. 182 Issue 3 Pg. 539-46 (Jul 01 1989) ISSN: 0014-2956 [Print] England
• PMID: 2473899 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Heat-Shock Proteins
• RNA, Messenger
• Poly A
• RNA
• DNA

Topics

• Amino Acid Sequence
• Base Sequence
• Cell Nucleus (metabolism)
• Chlamydomonas (genetics)
• Chloroplasts (metabolism)
• Cloning, Molecular
• DNA (isolation & purification)
• Genetic Code
• Heat-Shock Proteins (genetics, isolation & purification, metabolism)
• Molecular Sequence Data
• Nucleic Acid Hybridization
• Plasmids
• Poly A (isolation & purification)
• RNA (isolation & purification)
• RNA, Messenger (isolation & purification)
• Transcription, Genetic