N‐TERMINAL AMINO ACID SEQUENCE OF PROALBUMIN FROM INBRED BUFFALO RATS

Abstract

The sequence of radioactively labelled amino acids at the N‐terminus of proalbumin was determined by automated Edman‐degradation. [3H] Valine, [3H] phenylalanine or [14Carginine was incorporated into protein in vivo for a time period of 10 min after injection. Since albumin remains unlabelled during this time period (Urban et al., 1976), separation of proalbumin and albumin was not required for this work. Hence, compared to previous methods, a shorter purification procedure could be used which increased the yield of anti‐albumin‐precipitable protein and reduced the risk of proteolysis. Microsomes were prepared from livers removed 10 min after injection of the radioactively labelled amino acids. A buffer extract of the acetone‐dried powder from these microsomes was chromatographed on DEAE‐cellulose. All protein obtained after chromatography which could be precipitated with antiserum to serum albumin was isolated by immunoprecipitation and subsequent separation of the antigen‐antibody complex. The sequence of radioactive amino acids in this antigen preparation suggests that about 20–25% of proalbumin possessed at the N‐terminus the pentapeptide sequence X‐Val‐Phe‐Arg‐Arg‐ whereas 75–80% contained the hexapeptide sequence Arg‐X‐Val‐Phe‐Arg‐Arg‐.