Isolation of a Latimeria menadoensis heat shock protein 70 (Lmhsp70) that has all the features of an inducible gene and encodes a functional molecular chaperone.
Molecular chaperones facilitate the correct folding of other proteins, and heat shock proteins form one of the major classes of molecular chaperones. Heat shock protein 70 (Hsp70) has been extensively studied, and shown to be critically important for cellular protein homeostasis in almost all prokaryotic and eukaryotic systems studied to date. Since there have been very limited studies conducted on coelacanth chaperones, the main objective of this study was to genetically and biochemically characterize a coelacanth Hsp70. We have successfully isolated an Indonesian coelacanth (L. menadoensis) hsp70 gene, Lmhsp70, and found that it contained an intronless coding region and a potential upstream regulatory region. Lmhsp70 encoded a typical Hsp70 based on conserved structural and functional features, and the predicted upstream regulatory region was found to contain six potential promoter elements, and three potential heat shock elements (HSEs). The intronless nature of the coding region and the presence of HSEs suggested that Lmhsp70 was stress-inducible. Phylogenetic analyses provided further evidence that Lmhsp70 was probably inducible, and that it branched as a clade intermediate between bony fish and tetrapods. Recombinant LmHsp70 was successfully overproduced, purified and found to be functional using ATPase activity assays. Taken together, these data provide evidence for the first time that the coelacanth encodes a functional molecular chaperone system.
Modisakeng
Keoagile W
KW
Biomedical Biotechnology Research Unit, Department of Biochemistry, Microbiology and Biotechnology, Rhodes University, Grahamstown, South Africa.
Jiwaji
Meesbah
M
Pesce
Eva-Rachele
ER
Robert
Jacques
J
Amemiya
Chris T
CT
Dorrington
Rosemary A
RA
Blatch
Gregory L
GL
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D043344:Chordata / Q000235:genetics*; D022202:Chromosomes, Artificial, Bacterial / Q000235:genetics; D020451:Contig Mapping; D004926:Escherichia coli; D005786:Gene Expression Regulation; D018840:HSP70 Heat-Shock Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D010802:Phylogeny; D011401:Promoter Regions, Genetic / Q000235:genetics; D017433:Protein Structure, Secondary
D018840:HSP70 Heat-Shock Proteins; D000251:Adenosine Triphosphatases