Isolation of the V-ATPase A and c subunit cDNAs from mosquito midgut and Malpighian tubules.

摘要

Using conserved amino acid sequences for the design of oligonucleotide primers, cDNA clones for 2 subunits of the V-ATPase were isolated from the midgut and Malpighian tubules of Aedes aegypti larvae. The 3.1 kb cDNA of the A subunit of the peripheral catalytic V1 sector codes for a protein of 68.6 kDa. The protein contains conserved motifs, including an ATP/GTP binding site, found in all other A subunits. Southern analysis using the A subunit as a probe suggests the presence of only a single copyof the gene in A. aegypti. The 0.85 kb cDNA of the c subunit of the membrane H+ conducting V0 sector codes for a protein of 16.0 kDa. This protein has 4 transmembrane domains and contains a conserved glutamic acid that serves as the binding site for dicyclohexylcarbodiimide. Southern analysis using the c subunit as a probe suggests the presence of >1 related gene in the genome of A. aegypti. Pileup analysis of various A and c subunits shows that these subunits fall into distinct clusters, including onein which all arthropod proteins are clustered. The EMBL/GenBank/DDBJ accession numbers for the new nucleotide sequences are AF008922 (A subunit) and AF008924 (c subunit).