Characterization of cDNA clones for the beta subunit of pyruvate dehydrogenase from Ascaris suum

Abstract

The pyruvate dehydrogenase complex occupies a unique position in the anaerobic mitochondrial metabolism of the parasitic nematode Ascaris suum. This paper describes cDNA clones for the β subunit of the pyruvate dehydrogenase component of this complex. A cDNA library has been constructed in λgt11 from poly(A) + RNA isolated from adult ascarid body wall muscle. The library was screened with antiserum prepared against the β subunit of pyruvate dehydrogenase. Full-length clones of 1.2 kb have been characterized. The first 15 amino acids determined from the purified protein match exactly those predicted from the cDNA sequence. The deduced protein sequence contains a 26-amino-acid presequence that has characteristics of mitochondrial targeting sequences. The mature protein is predicted to contain 334 amino acids and is 62% identical to the predicted sequence of the corresponding human subunit [10]. Full-length in vitro transcripts have been translated in vitro to yield a 39-kDa polypeptide consistent with the open reading frame present in the cDNA sequence. The 90 nucleotides at the 3′ end of the cDNA sequence have the potential to form a cruciform structure that may play a role in the synthesis of the enzyme.