Enzymatic cleavage and HPLC peptide mapping of proteins

被引：22

作者：

Williams, KR

Stone, KL

机构：

[1] YALE UNIV,WM KECK FDN BIOTECHNOL RESOURCE LAB,NEW HAVEN,CT 06536

[2] YALE UNIV,HOWARD HUGHES MED INST,BIOPOLYMER FACIL,NEW HAVEN,CT 06536

来源：

MOLECULAR BIOTECHNOLOGY | 1997年 / 8卷 / 02期

关键词：

SDS-PAGE; HPLC; in-gel digestion; comparative HPLC peptide mapping; pepsin; trypsin; chymotrypsin; lysyl endopeptidase; protease V8;

D O I：

10.1007/BF02752260

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

Detailed procedures are described for successfully digesting reasonably small quantities (i.e., usually >10 pmol) of proteins with a variety of proteases and for then isolating the resulting peptides by reverse-phase HPLC. Since sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) appears to be the current method of choice for final purification of proteins for structural analysis, special attention is given to carrying out in-gel proteolytic digests on SDS-PAGE-separated proteins that have usually been stained with Coomassie Blue. A compilation of data from nearly 200 ''unknown'' samples is used to help provide realistic expectations with respect to the results that are likely to be obtained from carrying out in-gel proteolytic digests on large numbers of proteins.

引用

页码：155 / 167

页数：13

共 20 条

[1] INTERNAL AMINO-ACID SEQUENCE-ANALYSIS OF PROTEINS SEPARATED BY ONE-DIMENSIONAL OR TWO-DIMENSIONAL GEL-ELECTROPHORESIS AFTER INSITU PROTEASE DIGESTION ON NITROCELLULOSE [J].