Wang Sheng, Liu Ting, Zhang Luyan, Chen Gang
School of Pharmacy and Department of Chemistry, Fudan University, Shanghai 200032, China.
J Proteome Res. 2008 Nov;7(11):5049-54. doi: 10.1021/pr800476s. Epub 2008 Oct 18.
Infrared (IR) radiation was employed to enhance the efficiency of chymotryptic proteolysis for peptide mapping in this work. Protein solutions containing chymotrypsin in sealed transparent Eppendorf tubes were allowed to digest under an IR lamp at 37 degrees C. BSA and cytochrome c (Cyt- c) were digested by IR-assisted chymotryptic proteolysis to demonstrate the feasibility and performance of the novel digestion approach and the digestion time was significantly reduced to 5 min. The obtained digests were further identified by MALDI-TOF MS with the sequence coverages that were comparable to those obtained by using conventional in-solution digestion. The suitability of IR-assisted chymotryptic proteolysis to complex proteins was demonstrated by digesting human serum. The present proteolysis strategy is simple and efficient, offering great promise for high-throughput protein identification.
在本研究中，采用红外（IR）辐射提高胰凝乳蛋白酶解的效率以进行肽图谱分析。将含有胰凝乳蛋白酶的蛋白质溶液置于密封的透明艾本德管中，在37摄氏度的红外灯下进行消化。通过红外辅助胰凝乳蛋白酶解对牛血清白蛋白（BSA）和细胞色素c（Cyt-c）进行消化，以证明这种新型消化方法的可行性和性能，消化时间显著缩短至5分钟。通过基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）进一步鉴定所获得的消化产物，其序列覆盖率与使用传统溶液内消化获得的序列覆盖率相当。通过对人血清进行消化，证明了红外辅助胰凝乳蛋白酶解对复杂蛋白质的适用性。本蛋白水解策略简单高效，为高通量蛋白质鉴定提供了广阔前景。
