CHARACTERIZATION OF A DEVELOPMENTALLY RELATED POLYPEPTIDE WITH GLUTELIN SOLUBILITY CHARACTERISTICS FROM LUPINUS-ALBUS L

Proteins from Lupinus albus L. cv. Rio Maior seeds were fractionated a ccording to solubility criteria. Patterns of concanavalin A (ConA)-bin ding polypeptides from the different classes, albumins, globulins, glu telins and prolamins, were established by sodium dodecyl sulfate-polya crylamide gel electrophoresis. Two bands of apparent molecular masses of 29 and 23.5 kDa with glutelin solubility characteristics bound the lectin. The 23.5-kDa band was separated by two-dimensional electrophor esis into two components: one glycosylated and heterogeneous with an i soelectric point of approx. 10 (designated as G23) and another, not de tected with ConA, precipitating in the first dimension. The amino acid and hexosamine analysis of G23 showed that it is particularly rich in Gly (11.2%), Glx (10.0%), Ser (9.0%), Leu (8.2%), Asx (7.5%), and Pro (6.7%) and that it has a considerable content of the sulphur-containi ng amino acids Met (2.0%) and Cys (5.8%) and contains glucosamine. The determined N-terminal amino acid sequence of G23 was: (1)KG(R)V(5)KGT GD(10)(T)PXXV(15)XLY(N)R(20)T, and this had no significant similarity to any of the amino acid sequences contained in the data bank SWISS-PR OT 26. The glycoprotein G23 was completely deglycosylated with peptide -N-glycosidase F, yielding a homogeneous 21-kDa polypeptide composed o f approximately 191 amino acids. The structures of the major N-linked neutral oligosaccharides of G23, determined by exoglycosidase sequenci ng, were as follows: Man alpha 2Man alpha 6(Man alpha 3) Man alpha 6(M an alpha 2Man alpha 2Man alpha 3)Man beta 4GlcNAc beta 4 GlcNAc (13%); +/- Man alpha 2Man alpha 6(Man alpha 3)Man alpha 6(+/- Man alpha 2 Ma n alpha 2 Man alpha 3) Man beta 4GlcNAc beta 4GlcNAc (29%); Man alpha 6(Man alpha 3) Man alpha 6(Man alpha 2Man alpha 3)Man beta 4GlcNAc bet a 4GlcNAc (13%); Man alpha 6(Man alpha 3)Man alpha 6(Man alpha 3)Man b eta 4GlcNAc beta 4Glc-NAc (16%); Man alpha 6(Man alpha 3)(Xyl beta 2)M an beta 4GlcNAc beta 4Glc-NAc (28%). Changes in G23 abundance during s eed development, germination and seedling growth were monitored with a specific antibody. The glycoprotein G23 started to accumulate appreci ably during seed formation between the 40th and the 50th days after an thesis and was detected following seed imbibition, until the 9th day i n cotyledons, the 2nd day in roots and the 4th day in hypocotyls and l eaves.