Establishment of the human reflex tear two-dimensional polyacrylamide gel electrophoresis reference map: new proteins of potential diagnostic value

Mark P. Molloy, Shirley Bolis, Ben R. Herbert, Keli Ou, Margaret I. Tyler, Derek D. Van Dyk, Mark D. P. Willcox, Andrew A. Gooley, Keith L. Williams, Carol A. Morris, Bradley J. Walsh*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

97 Citations (Scopus)

Abstract

To understand the changes in protein expression associated with various physiological states as well as the development of pathological eye disease, we have begun to map the protein components of normal human reflex tears. An analytical reference map of normal human reflex tears was created using two- dimensional polyacrylamide gel electrophoresis (2-D PAGE) with pH 3.5-10 immobilized pH gradients (IPGs). Micropreparatively loaded gels were transferred to polyvinylidene difluoride (PVDF) and analysed by a combination of N-terminal sequence tagging and amino acid compositional analysis. Thirty spots were sequence tagged, resulting in identification of six different proteins (lipocalin, lysozyme, lactotransferrin, zinc-α-2 glycoprotein, cystatin S, cystatin SN) that matched to entries in the SWISS-PROT database. A group of N-terminally blocked proteins was clearly identified from SWISS-PROT by amino acid analysis, isoelectric point (pI) and molecular weight (M(r)). A number of highly expressed protein components remain unidentified despite being subjected to amino acid analysis and Edman sequencing. A majority of the abundant proteins showed varying degrees of charge heterogeneity attributed to post-translational processing such as glycosylation and N-terminal truncation. We have identified a previously undescribed protein that we have named lacryglobin. This protein displays strong homology with mammaglobin, a protein overexpressed in breast cancer. The discovery of this homologue in tears offers the potential for disease diagnosis by screening tear fluid proteins.

Original languageEnglish
Pages (from-to)2811-2815
Number of pages5
JournalElectrophoresis
Volume18
Issue number15
DOIs
Publication statusPublished - Dec 1997

Keywords

  • amino acid analysis
  • cancer
  • lacryglobin
  • protcome
  • tag Sequencing
  • tears
  • two-dimensional polyacrylamide gel electrophoresis

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