Fisichella S, Foti S, Maccarrone G, Saletti R
Dipartimento di Scienze Chimiche, Università di Catania, Italy.
J Chromatogr A. 1995 Feb 17;693(1):33-44. doi: 10.1016/0021-9673(94)01042-d.
The determination of the tryptic peptide mapping of sequence 299-585 (cyanogen bromide fragment A) of human serum albumin (HSA) by chemical and enzymatic cleavages and combined use of HPLC and FAB-MS is described. Reduction and carboxymethylation of A gave four subfragments which were separated by HPLC and digested with trypsin. Tryptic fragments were separated by HPLC and identified by FAB-MS. A total coverage of about 95% of the entire sequence was obtained. Tryptic fragments not identified include mostly single amino acids and very hydrophilic peptides which were absent in the chromatograms. The high reproducibility of the experiments and the satisfactory yield of the tryptic fragments identified demonstrate the great potential of the combined use of HPLC separation and FAB-MS analysis for the structural investigation of HSA.
本文描述了通过化学和酶切、高效液相色谱（HPLC）和快原子轰击质谱（FAB-MS）联用，对人血清白蛋白（HSA）序列299 - 585（溴化氰片段A）进行胰蛋白酶肽图谱分析的过程。对片段A进行还原和羧甲基化处理后得到四个亚片段，它们通过HPLC分离，并用胰蛋白酶消化。胰蛋白酶消化片段通过HPLC分离，并由FAB-MS鉴定。整个序列的总覆盖率约为95%。未鉴定出的胰蛋白酶片段主要包括单个氨基酸和色谱图中未出现的极亲水肽段。实验的高重现性以及所鉴定胰蛋白酶片段的满意产率证明了HPLC分离和FAB-MS分析联用在HSA结构研究中的巨大潜力。
