An analysis of protein abundance suppression in data dependent liquid chromatography and tandem mass spectrometry with tryptic peptide mixtures of five known proteins

Eur. J. Mass Spectrom. 11, 575–580 (2005)
DOI: 10.1255/ejms.776

An analysis of protein abundance suppression in data dependent liquid chromatography and tandem mass spectrometry with tryptic peptide mixtures of five known proteins

Wei Sun,^* Shuzhen Wu, Xiaorong Wang, Dexian Zheng, Youhe Gao^*
Proteomics Research Center National Key Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Peking Union Medical College/Chinese Academy of Medical Sciences, 100005 Beijing, People’s Republic of China. E-mail: gaoyouhe@pumc.edu.cn or sunwei1018@hotmail.com

ABSTRACT:

Reverse phase liquid chromatography (RPLC) has been widely used in proteomics research for peptide separation. When protein samples are separated by RPLC and identified with electrospray ion trap mass spectrometry (ESI-MS), the signals of high-abundance proteins may suppress those of low-abundance proteins, a phenomenon known as abundance suppression. To what degree the abundance suppression correlates to the number of tryptic peptides in the high-abundance proteins has not been carefully investigated. We tried to answer this question by studying the mixtures digested from five known proteins. The numbers of identified tryptic peptides (longer than five amino acids) of the five proteins ranged from 12 to 47. Four different peptide mixtures with 10- to 100-fold abundance differences of five known proteins were separated by RPLC and identified by ESI-MS. Our results showed that abundance suppression was related to the tryptic peptide numbers in the high-abundance protein. Within a 100-fold protein abundance difference range, tryptic peptide number in the low-abundance proteins could be suppressed up to seven times by high-abundance proteins. The procedure we suggest here can help to identify low-abundance proteins co-purified with their high-abundance binding protein. The result can also help to identify specific high-abundance proteins for removal by immunoaffinity.

Keywords: protein identification, abundance suppression, reverse phase liquid chromatography, electrospray ion trap mass spectrometry

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