title: Glutathione S-transferase and S-crystallins of cephalopods: evolution from active enzyme to lens-refractive proteins.
abstract: Our previous studies have shown that the S-crystallins of cephalopod (Ommastrephes sloani pacificus) eye lenses comprise a family of at least ten members which are evolutionarily related to glutathione S-transferase (GST, EC 2.5.1.18). Here we show by cDNA cloning that there are at least 24 different S-crystallins that are 46-99% identical to each other by amino acid sequence in the squid Loligo opalescens. In each species, all but one S-crystallin (SL11 in O. pacificus and Lops4 in L. opalescens) examined has an inserted central peptide of variable length and sequence. cDNA expression studies conducted in Escherichia coli showed that squid GST (which is expressed little in the lens) has very high enzymatic activity using 1-chloro-2, 4-dinitrobenzene (CDNB) as a substrate; by contrast, SL20-1 of O. pacificus and Lops12 of L. opalescens (which are encoded by abundant lens mRNAs) have no GST activity. Interestingly, SL11 and Lops4 have some enzymatic activity with the CDNB substrate. Site-specific mutations at Y7 or W38, both residues essential for activity of vertebrate GSTs, or insertion of the central peptide present in the inactive SL20-1, reduced the specific activity of squid GST by 30- to 100-fold. These data indicate that the S-crystallins consist of a family of enzymatically inactive proteins (when using CDNB as a substrate) which is considerably larger than previously believed and that GST activity was lost by gradual drift in sequence as well as by insertion of an extra peptide by exon shuffling. The results are also consistent with the idea that SL11 and Lops4 are orthologous crystallins representing the first descendants of the ancestral GST gene in the pathway which gave rise to the extensive S-crystallin family of lens proteins.
authors.affiliation: Laboratory of Molecular and Developmental Biology, National Eye Institute, National Institutes of Health, Bethesda, MD 20892-2730, USA.
authors.forename: S I
authors.initials: SI
authors.lastname: Tomarev
authors.forename: S
authors.initials: S
authors.lastname: Chung
authors.forename: J
authors.initials: J
authors.lastname: Piatigorsky
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D003459:Crystallins / Q000235:genetics*; D049832:Decapodiformes / Q000235:genetics* / Q000378:metabolism; D004789:Enzyme Activation; D019143:Evolution, Molecular; D005982:Glutathione Transferase / Q000235:genetics*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D016415:Sequence Alignment
chemical_list: D003459:Crystallins; D005982:Glutathione Transferase