Work

Microsequence and mass spectral analysis of nonspecific cross-reacting antigen 160, a CD15-positive neutrophil membrane glycoprotein. Demonstration of identity with biliary glycoprotein

Year: 1993

Type: article

Abstract: Sequence information was obtained from low picomole amounts of nonspecific cross-reacting antigen (NCA) 160 (M(r) 160,000), a granulocyte membrane glycoprotein. Following affinity purification and SDS-polyacrylamide gel electrophoresis, the protein was electrotransferred to nitrocellulose, digested with trypsin, and the peptides were isolated using capillary reversed-phase liquid chromatography. Analysis of these peptides by Edman microsequencing and mass spectrometry established that NCA-160 was identical to biliary glycoprotein I, a protein that we previously cloned from a human colon library (1). NCA-160 from human granulocytes is a CD15-positive glycoprotein belonging to the carcinoembryonic antigen family and possesses putative transmembrane and cytoplasmic domains. Previous efforts to characterize this antigen at the protein level were hampered by a blocked NH2 terminus. In this study, we confirmed 20% of the deduced amino acid sequence starting with approximately 50 pmol of sample. Carbohydrate structural data is also presented on a single N-linked oligosaccharide moiety located in the A' domain. The capillary high performance liquid chromatography techniques used here, as well as mass spectrometry, were essential for high sensitivity analysis of the blotted, digested glycoprotein.

Source: Journal of Biological Chemistry

DOI: 10.1016/s0021-9258(19)38610-7

Authors: A.M. Mahrenholz, C.H. Yeh, J.E. Shively, S.A. Hefta

Institutions: City Of Hope National Medical Center, City of Hope, Beckman Research Institute

Language: English

Topic: Advanced Proteomics Techniques and Applications

Subfield: Spectroscopy

Field: Chemistry

Domain: Physical Sciences

SDG: Good health and well-being

Open Access status: hybrid

Estimated APC paid: $3,430