Cloning of the esterase-5 locus from Drosophila pseudoobscura and comparison with its homologue in D. melanogaster.
A clone of the esterase-5 (Est-5) gene from Drosophila pseudoobscura has been isolated by hybridization to the cloned Est-6 gene of D. melanogaster. Southern analysis and sequencing of the cloned DNA revealed three regions of similarity to Est-6 that have been tentatively identified as genes, Est-5A, Est-5B, and Est-5C. Introduction of each of the three genes separately into D. melanogaster by P-element transformation has demonstrated that Est-5B encodes an enzyme with the same physical properties as EST 5 in D. pseudoobscura. Sequence analysis indicates that Est-5B encodes a 545-amino-acid protein and is composed of two exons separated by a 55-bp intron in the same position as the 51-bp intron in Est-6. Comparison of the Est-5B coding region with that of Est-6 reveals an overall similarity (73% at both the nucleotide and amino acid levels) that is substantially lower than that for other genes sequenced in both of these species. Total nucleotide and nonsynonymous site differences between Est-6 and Est-5B are more abundant in the second exon than in the first, suggesting differential effects of selection or mutation on these two exons. Comparisons of the 5'-flanking DNA of Est-5B and Est-6 reveal four short conserved sequence elements, but the remaining upstream sequences show no significant similarity. Conservation in the 3'-flanking DNA is limited to the presence of two polyadenylation sites that may correlate with the existence of two transcripts from both Est-5B and Est-6. The patterns of nucleotide substitutions and amino acid replacements between Est-5B and Est-6 are consistent with the hypothesis that mutation and genetic drift are responsible for the differences between these two genes.
Brady
J P
JP
Department of Biology, Indiana University.
Richmond
R C
RC
Oakeshott
J G
JG
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D043182:Carboxylesterase; D002265:Carboxylic Ester Hydrolases / Q000235:genetics; D003001:Cloning, Molecular; D004251:DNA Transposable Elements; D004330:Drosophila / Q000201:enzymology / Q000235:genetics*; D029721:Drosophila Proteins; D004331:Drosophila melanogaster / Q000201:enzymology / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D004950:Esterases / Q000235:genetics*; D005796:Genes; D008969:Molecular Sequence Data; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D014170:Transformation, Genetic
D004251:DNA Transposable Elements; D029721:Drosophila Proteins; D004950:Esterases; D002265:Carboxylic Ester Hydrolases; D043182:Carboxylesterase; C468266:Est-6 protein, Drosophila