cDNA cloning, expression, and characterization of an arylphorin-like hexameric storage protein, AgeHex2, from the mulberry longicorn beetle, Apriona germari.
An arylphorin-like hexameric storage protein, AgeHex2, cDNA was cloned from the mulberry longicorn beetle, Apriona germari (Coleoptera, Cerambycidae), larval cDNA library. The complete cDNA sequence of AgeHex2 is comprised of 2,088 bp encoding 696 amino acid residues. The AgeHex2 had four potential N-glycosylation sites. The AgeHex2 contained the highly conserved two larval storage protein signature motifs. The deduced protein sequence of AgeHex2 showed high homology with A. germari hexamerin1 (51% amino acid identity), Tenebrio molitor hexamerin2 (49% amino acid identity), T. molitor early-staged encapsulation inducing protein (43% amino acid identity), and Leptinotarsa decemlineata diapause protein1 (43% amino acid identity). Phylogenetic analysis further confirmed the AgeHex2 is more closely related to coleopteran hexamerins than to the other insect storage proteins. Northern blot analysis confirmed that the AgeHex2 showed fat body-specific expression. The cDNA encoding AgeHex2 was expressed as a 75-kDa protein in the baculovirus-infected insect cells. Furthermore, N-glycosylation of the recombinant AgeHex2 was revealed by tunicamycin to the recombinant virus-infected Sf9 cells, demonstrating that the AgeHex2 is N-glycosylated. Western blot analysis using the polyclonal antiserum against recombinant AgeHex2 indicated that the AgeHex2 corresponds to a 75-kDa storage protein present in the A. germari larval hemolymph.
Kim
Seong Ryul
SR
College of Natural Resources and Life Science, Dong-A University, Busan, Korea.
Lee
Kwang Sik
KS
Yoon
Hyung Joo
HJ
Park
Nam Sook
NS
Lee
Sang Mong
SM
Je
Yeon Ho
YH
Jin
Byung Rae
BR
Sohn
Hung Dae
HD
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D001517:Coleoptera / Q000235:genetics* / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D006023:Glycoproteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics*; D006031:Glycosylation; D019476:Insect Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics*; D007814:Larva / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D010802:Phylogeny; D020836:Protein Structure, Quaternary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D016415:Sequence Alignment; D014018:Tissue Distribution; D014158:Transcription, Genetic
D000596:Amino Acids; D018076:DNA, Complementary; D006023:Glycoproteins; D019476:Insect Proteins; D011994:Recombinant Proteins; C041137:storage proteins, Insecta