ISOLATION, CHARACTERIZATION BY MASS-SPECTROMETRY AND PARTIAL AMINO-ACID SEQUENCING OF AVENINS

Avenins were extracted with 70% (v/v) ethanol and purified by IE-HPLC followed by RP-HPLC. Ten major and one minor avenins were isolated and characterised. The amino acid compositions of the avenins were simila r to those of the alcohol-extractable seed storage proteins of other c ereals. They were rich in proline and glutamine and poor in basic amin o acids. The exact number of cysteine residues in the avenins was dete rmined by ESMS before and after reduction and alkylation. Free SH grou ps were not found in Avn-9 and Avn-3, and four disulphide bonds are th erefore expected. The M(r)s of the avenins ranged between 20 and 28k a s determined by mass spectrometry. A difference between the M(r) calcu lated from the amino acid sequence of Avn-9 and the M(r) measured by E SMS was indicative of an erroneous sequence, and mass spectrometric pe ptide mapping in combination with Edman degradation led to the identif ication of three sequence errors. Five of the avenins were N-terminall y blocked. The N-terminal sequences of the six other avenins revealed unique highly homologous regions and exhibited short tandem repeats.