An application of diagonal electrophoresis to the selective purification of serine phosphate peptides. Serine phosphate peptides from ovalbumin.
A diagonal-electrophoresis method for the selective purification of serine phosphate peptides was applied to tryptic, chymotryptic and peptic digests of oxidized ovalbumin. This method is based on the release of the phosphate group bound to serine by treatment with alkaline phosphatase on paper. The identified serine phosphate peptides were purified by paper electrophoresis at pH6.5 and 2.0, dephosphorylation with bacterial alkaline phosphatase, and paper electrophoresis at pH2.0 again, in that order. The presence of two groups of serine phosphate peptides was apparent from the amino acid composition. One group contained no lysine, cysteic acid, proline, leucine or isoleucine (sequence 1) and the other had all those amino acids (sequence 2). Further degradation with subtilisin of those peptides and ;dansyl'-Edman sequence analysis established their partial sequences. The proposed sequences are as follows (with ;SerP' representing serine phosphate): sequence 1, -Ala-Gly-Arg-Glu-Val-Val-Gly-SerP-Ala-Glu-Ala-Gly-Asp-Val-Ala-Ala-Ser-(Val,Glx(2),Ser,Phe)-Arg-; sequence 2, -Asp-Lys-Leu-Pro-Gly-Phe-Gly-Asp-SerP-Ile-Glx-Ala-Glx-CySO(3)H-Gly-(Thr,Ser,Val)-(Asp,His,Val)-. The partial sequence of one of the phosphopeptides, Asp-(Glu,Ile,SerP), reported by Flavin (1954) was used to establish the proposed sequence 2.
D000469:Alkaline Phosphatase; D000595:Amino Acid Sequence; D002918:Chymotrypsin; D004586:Electrophoresis; D007532:Isoleucine / Q000032:analysis; D007930:Leucine / Q000032:analysis; D008239:Lysine / Q000032:analysis; D010047:Ovalbumin / Q000032:analysis*; D010434:Pepsin A; D010455:Peptides / Q000302:isolation & purification*; D011392:Proline / Q000032:analysis; D012694:Serine / Q000302:isolation & purification*; D014357:Trypsin
D010455:Peptides; D007532:Isoleucine; D012694:Serine; D010047:Ovalbumin; D011392:Proline; D000469:Alkaline Phosphatase; D002918:Chymotrypsin; D014357:Trypsin; D010434:Pepsin A; D007930:Leucine; D008239:Lysine