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Sequence Analysis of Frog Rho-Crystallin by cDNA Cloning and Sequencing: A Member of the Aldo-Keto Reductase Family

Sequence Analysis of Frog Rho-Crystallin by cDNA Cloning and Sequencing: A Member of the Aldo-Keto Reductase Family

S F Lu, F M Pan, S H Chiou

Biochem Biophys Res Commun. 1995 Sep 25;214(3):1079-88.

PMID: 7575513

Abstract:

rho-Crystallin is a major enzyme crystallin present in the lenses of amphibian species with a blocked amino terminus. In order to facilitate the determination of the primary sequence of this taxon-specific crystallin, cDNA mixture was synthesized from the poly(A)+mRNA of bullfrog eye lenses. cDNAs encoding rho-crystallin were then amplified by polymerase chain reaction (PCR) using a new protocol of Rapid Amplification of cDNA Ends (RACE). PCR-amplified product corresponding to rho-crystallin was obtained, which was then subcloned into pUC18 vector and then transformed into E. coli strain JM109. Plasmids purified from the positive clones were prepared for nucleotide sequencing by the automatic fluorescence-based dideoxynucleotide chain-termination method. Sequencing more than 15 clones containing DNA inserts coding for rho-crystallin constructed only one unique and complete full-length reading frame of 975 base pairs covering a deduced protein sequence of 324 amino acids including the universal initiating methionine. It shows 96, 59, 46 and 37 percent sequence similarity to another rho-crystallin from European common frog, bovine prostaglandin-F synthase, human aldose reductase and human aldehyde reductase, respectively, revealing the close relationship between rho-crystallins from related amphibian species and its possible evolutionary relatedness with various aldo-keto reductases. In this study a phylogenetic tree for rho-crystallin and related enzymes is constructed based on multiple-sequence alignment program using a combination of distance matrix and approximate parsimony methods. We have thus established the remote phylogenetic relationship between rho-crystallin and some aldehyde/aldose reductases, which may provide a possible link for the recruitment of this crystallin from detoxification-related enzymes and its physiological role in maintaining a transparent and clear lens.

Chemicals Related in the Paper:

Catalog Number	Product Name	Structure	CAS Number	Price
AP55976959	Prostaglandin F Synthase human		55976-95-9	Price

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