Cloning and expression of an insect Ca(2+)-ATPase from Heliothis virescens.
A complementary DNA for the Tobacco Budworm, Heliothis virescens, sarco(endo)plasmic reticulum-type Ca(2+)-ATPase (HVSERCA) has been cloned and sequenced. cDNA fragments of adult rabbit fast-twitch muscle Ca(2+)-ATPase (SERCA1a) were used as heterologous probes to isolate a partial cDNA clone coding for a protein with high homology to the Ca(2+)-ATPase from Drosophila melanogaster (DRSERCA) and vertebrate ER/SR Ca2+ pumps. The entire cDNA clone contains an ORF encoding a protein of 1000 amino acids which shares the characteristic motifs of a P-type ATPase. HVSERCA shares 89% identity with DRSERCA, 80% identity with the Artemia Ca(2+)-ATPase and 72% identity with avian and mammalian SERCAs. An insect Ca(2+)-ATPase-specific polyclonal antiserum has been raised against a fusion protein containing sequence from the cytoplasmic domain of HVSERCA. Heterologous expression of the insect pump in COS-7 cells has been demonstrated by immunocytochemistry and the reticular pattern of staining is consistent with an ER localisation. However, the expressed enzyme from COS-7 cells does not appear to be active.
Lockyer
P J
PJ
Division of Biochemistry and Molecular Biology, School of Biological Sciences, University of Southampton, UK.
Puente
E
E
Windass
J
J
Earley
F
F
East
J M
JM
Lee
A G
AG
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D000252:Calcium-Transporting ATPases / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary; D008969:Molecular Sequence Data; D009036:Moths / Q000201:enzymology / Q000235:genetics*; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D000252:Calcium-Transporting ATPases