title: Molecular characterization and expression of a gene encoding cytosolic Hsp90 from Pennisetum glaucum and its role in abiotic stress adaptation.
abstract: Heat shock protein 90 (Hsp90) is an abundant and highly conserved molecular chaperone that is essential for viability in eukaryotes. They have a crucial role in the folding of a set of proteins involved in the regulation of many essential cellular pathways and also re-folding of stress-denatured polypeptides. However, their exact function is still not clearly elucidated. In this study the full-length cDNA encoding for Hsp90 polypeptide and its corresponding gene was isolated from Pennisetum glaucum (designated PgHsp90). PgHsp90 cDNA encoded for a polypeptide of 698 amino acids with a predicted molecular mass of 80.3kDa and shared a high sequence homology (97-81%) to other plant cytosolic Hsp90s and shared less sequence homology (40-45%) to organelle and endoplasmic reticulum specific Hsp90 isoforms. A deduced amino acid sequence possessed three structural domains: N-terminus (1-211) ATP binding domain, middle (281-540) client protein interacting domain and C-terminus (541-698) dimerization domain; the N-terminus and middle domain is linked by a charged linker domain (212-280). It possesses the five-conserved amino acid signature sequence motifs characteristic of the Hsp90 family and a C-terminus MEEVD penta-peptide characteristic of the cytosolic Hsp90 isoform. The predicted quaternary architecture generated for PgHsp90 through molecular modeling was globally akin to that of yeast Hsp90. The PgHsp90 gene consists of 3 exons and 2 introns. The position and phasing of these introns were conserved in other plant cytosolic Hsp90 genes. Recombinant PgHsp90 protein was expressed in E. coli and purified to homogeneity, which possessed in vitro chaperone activity. E. coli expressing PgHsp90 protein showed enhanced tolerance to heat, salt and dehydration stresses. The quantitative up-regulation of PgHsp90 gene expression positively correlates in response to different stresses to meet the additional demand for protein folding support. Cumulatively, the in vivo and in vitro experiments indicated that PgHsp90 plays an adaptive or protective role to counter the stress induced protein damage.
authors.affiliation: Plant Molecular Biology Group, International Centre for Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, New Delhi 110 067, India.
authors.forename: Palakolanu Sudhakar
authors.initials: PS
authors.lastname: Reddy
authors.forename: V
authors.initials: V
authors.lastname: Thirulogachandar
authors.forename: C S
authors.initials: CS
authors.lastname: Vaishnavi
authors.forename: A
authors.initials: A
authors.lastname: Aakrati
authors.forename: Sudhir K
authors.initials: SK
authors.lastname: Sopory
authors.forename: Malireddy K
authors.initials: MK
authors.lastname: Reddy
mesh_terms: D000222:Adaptation, Physiological / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D017124:Conserved Sequence / Q000235:genetics; D005091:Exons; D015870:Gene Expression; D018841:HSP90 Heat-Shock Proteins / Q000235:genetics* / Q000378:metabolism; D007438:Introns; D008969:Molecular Sequence Data; D031782:Pennisetum / Q000235:genetics*; D011485:Protein Binding / Q000235:genetics; D017510:Protein Folding; D020836:Protein Structure, Quaternary; D017385:Sequence Homology; D013312:Stress, Physiological / Q000235:genetics*; D015854:Up-Regulation
chemical_list: D018841:HSP90 Heat-Shock Proteins