Proteome Technology

Identification of yeast proteins from two-dimensional gels: Working out spot cross-contamination

  1. Dr. Kenneth C. Parker1,*,
  2. James I. Garrels2,
  3. Wade Hines1,
  4. Erin M. Butler2,
  5. Andrew H. Z. McKee2,
  6. Dale Patterson1,
  7. Steve Martin1

Article first published online: 14 APR 2005

DOI: 10.1002/elps.1150191110

Issue

ELECTROPHORESIS

ELECTROPHORESIS

Volume 19, Issue 11, pages 1920–1932, August 1998

Additional Information

How to Cite

Parker, K. C., Garrels, J. I., Hines, W., Butler, E. M., McKee, A. H. Z., Patterson, D. and Martin, S. (1998), Identification of yeast proteins from two-dimensional gels: Working out spot cross-contamination. ELECTROPHORESIS, 19: 1920–1932. doi: 10.1002/elps.1150191110

Author Information

  1. 1

    PerSeptive Biosystems, Framingham, MA, USA

  2. 2

    Proteome Inc., Beverly, MA, USA

*500 Old Connecticut Path, Framingham, MA 01701, USA (Tel: +508-383-7426; Fax: +508-383-7883)

Publication History

  1. Issue published online: 14 APR 2005
  2. Article first published online: 14 APR 2005
  3. Manuscript Received: 7 MAY 1998

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Keywords:

  • Peptide mass fingerprinting;
  • Yeast;
  • Two-dimensional polyacrylamide gel electrophoresis;
  • Keratin

Abstract

With the complete sequence of the yeast genome now available, efforts by many laboratories are underway to identify each of the spots on two-dimensional (2-D) gels corresponding to the most abundant yeast proteins. The high mass accuracy now attainable using matrix assisted laser desorption/ionization (MALDI)-mass spectrometry equipped with delayed extraction simplifies the process of identification, such that many spots can be unambiguously identified in a short period of time merely by using peptide mass fingerprinting and generally available database matching programs. Although it is not always possible to match spots between gels run by different laboratories, proteins generally yield the same abundant proteolytic fragments when tryptic digestions are performed. Databases containing these signature peptides not only simplify the task of reidentifying proteins from different gels, but also make it possible to identify small amounts of cross-contaminating proteins from different spots, as well as common extraneous contaminants such as human keratins. In this paper, we present data on the identification of > 20 previously unreported yeast proteins from 2-D gels. Some novel proteins were identified from randomly analyzed spots. Focusing on 14 spots in a narrow-pH-range gel, we demonstrate how organizing peak-table data and peptide match-list data into databases enables the identification of a larger percentage of the peaks.

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