Abstract
We purified vitellin (ApVn) and vitellogenin (ApVg) from Antheraea pernyi by a combination of gel permeation chromatography, anion-exchange chromatography, and hydrophobic chromatography. Our results showed that the molecular size of the ApVg heavy chain determined by SDS-PAGE was approximately 210 kDa. ApVn and ApVg each consisted of only a large subunit, suggesting that ApVg should be classified in group 2 of the insect vitellogenin family. We analyzed the N-terminal amino acid sequence of ApVg and the terminal amino acid sequences of four lysyl endopeptidase-degraded fragments of ApVg. The nucleotide sequence was determined using overlapping cDNA fragments generated from reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE) reactions. The ApVg cDNA was 5720 nucleotides long and coded for the entire subunit, which consisted of 1778 amino acids. The molecular weight of the predicted polypeptide was 200, 000. There is no RXRR motif, which is the cleavage site between the small and large subunits in the previtellogenins of the silkworm, Bombyx mori, the mosquito, Aedes aegypti, the sawfly, Athalia rosae, and the boll weevil, Anthonomous grandis. Two polyserine regions were found in the deduced amino acid sequence.
