Use of peptide retention time prediction for protein identification by off-line reversed-phase HPLC-MALDI MS/MS
Use of peptide retention time prediction for protein identification by off-line reversed-phase HPLC-MALDI MS/MS
复制标题
DOI：
10.1021/ac060251b
复制
发表时间：
2006-09-01
影响因子：
7.4
通讯作者：
Wilkins, John A.
中科院分区：
文献类型：
--
作者：
Krokhin, Oleg V.;Ying, Stephen;Wilkins, John A.
开发了一种新的算法，序列特定保留计算器，用于预测胰蛋白酶多肽在C18,300埃孔径色谱柱上的反相高效液相分离过程中的保留时间。对于2000个(类似)多肽的测试库，相关系数高达0.98R-2值，对于各种实际样品，相关系数类似于0.95-0.97。该算法与基于质量(15ppm容差)和保留时间(0.66%乙腈/分钟梯度的2min容差)、MART标准的排除方案相结合，显著减少了用反相高效液相色谱分离的消化液进行完整MS/MS分析所需的仪器时间。尽管虚拟执行的MS/MS分析的数量减少了57%，但通过重新分析一组HPLC-MALDI MS/MS数据，在蛋白质鉴定方面没有损失，证实了这一点。
A new algorithm, sequence-specific retention calculator, was developed to predict retention time of tryptic peptides during RP HPLC fractionation on C18, 300-angstrom pore size columns. Correlations of up to similar to 0.98 R-2 value were obtained for a test library of (similar to) 2000 peptides and similar to 0.95-0.97 for a variety of real samples. The algorithm was applied in conjunction with an exclusion protocol based on mass ( 15 ppm tolerance) and retention time (2-min tolerance for 0.66% acetonitrile/min gradient), MART criteria to significantly reduce the instrument time required for complete MS/MS analysis of a digest separated by RP HPLC. This was confirmed by reanalyzing the set of HPLC-MALDI MS/MS data with no loss in protein identifications, despite the number of virtually executed MS/MS analyses being decreased by 57%.