Characterization of gamma-crystallin from a catfish: structural characterization of one major isoform with high methionine by cDNA sequencing.
gamma-Crystallin is the major and most abundant lens protein present in the eye lens of most teleostean fishes. To facilitate structural characterization of gamma-crystallins isolated from the lens of the catfishes (Clarias fuscus), a cDNA mixture was synthesized from the poly(A)+mRNA isolated from fresh eye lenses, and amplification by polymerase chain reaction (PCR) was adopted to obtain cDNAs encoding various gamma-crystallins. Plasmids of transformed E. coli strain JM109 containing amplified gamma-crystallin cDNAs were purified and prepared for nucleotide sequencing by the dideoxynucleotide chain-termination method. Sequencing more than five clones containing DNA inserts of 0.52 kb revealed the presence of one major isoform with a complete reading frame of 534 base pairs, covering a gamma-crystallin (gamma M1) with a deduced protein sequence of 177 amino acids excluding the initiating methionine. It was of interest to find that this crystallin of pI 9.1 contains a high-methionine content of 15.3% in contrast to those gamma-crystallins of low-methionine content from most mammalian lenses. Sequence comparisons of catfish gamma M1-crystallin with those published sequences of gamma-crystallins from carp, bovine and mouse lenses indicate that there is approx. an 82% sequence homology between the catfish and the carp species of piscine class whereas only 51-58% homology is found between mammals and the catfish. Moreover the differences in the hydropathy profiles for these two groups of gamma-crystallins, i.e. one with a high-methionine content from teleostean fishes and the other with a low-methionine content from mammalian species, reflect a distinct variance in the polarity distributions of surface amino acids in these crystallins.(ABSTRACT TRUNCATED AT 250 WORDS)
Pan
F M
FM
Institute of Biochemical Sciences, National Taiwan University, Academia Sinica, Taipei.
Chang
W C
WC
Lin
C H
CH
Hsu
A L
AL
Chiou
S H
SH
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002347:Carps; D002397:Catfishes / Q000502:physiology*; D002417:Cattle; D003459:Crystallins / Q000737:chemistry* / Q000235:genetics; D018076:DNA, Complementary / Q000096:biosynthesis; D007908:Lens, Crystalline / Q000737:chemistry*; D008715:Methionine / Q000737:chemistry; D051379:Mice; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D014867:Water / Q000737:chemistry
D003459:Crystallins; D018076:DNA, Complementary; D014867:Water; D008715:Methionine