Biosynthesis of human sialophorins and analysis of the polypeptide core
Biosynthesis was examined of sialophorin (formerly called gpL115) which is altered in the inherited immunodeficiency Wiskott-Aldrich syndrome. Sialophorin is greater than 50% carbohydrate, primarily O-linked units of sialic acid, galactose, and galactosamine. Pulse-labeling with (/sup 35/S)methionine and chase incubation established that sialophorin is synthesized in CEM lymphoblastoid cells as an Mr 62,000 precursor which is converted within 45 min to mature glycosylated sialophorin, a long-lived molecule. Experiments with tunicamycin and endoglycosidase H demonstrated that sialophorin contains N-linked carbohydrate (approximately two units per molecule) and is therefore an N,O-glycoprotein. Pulse-labeling of tunicamycin-treated CEM cells together with immunoprecipitation provided the means to isolate the (/sup 35/S)-methionine-labeled polypeptide core of sialophorin and determine its molecular weight (58,000). This datum allowed us to express the previously established composition on a per molecule basis and determine that sialophorin molecules contain approximately 520 amino acid residues and greater than or equal to 100 O-linked carbohydrate units. A recent study showed that various blood cells express sialophorin and that there are two molecular forms: lymphocyte/monocyte sialophorin and platelet/neutrophil sialophorin. Biosynthesis of the two forms was compared by using sialophorin of CEM cells and sialophorin of MOLT-4 cells (another lymphoblastoid line) as models for lymphocyte/monocyte sialophorin and platelet/neutrophil sialophorin, respectively. The time course of biosynthesis and the content of N units were found to be identical for the two sialophorin species. (/sup 35/S)Methionine-labeled polypeptide cores of CEM sialophorin and MOLT sialophorin were isolated and compared by electrophoresis, isoelectrofocusing, and a newly developed peptide mapping technique.
- Research Organization:
- The Children's Hospital, Boston, MA
- OSTI ID:
- 5842105
- Journal Information:
- Biochemistry; (United States), Vol. 26:13
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
GLUCOPROTEINS
BIOSYNTHESIS
CHEMICAL COMPOSITION
ELECTROPHORESIS
IMMUNOASSAY
METHIONINE
MOLECULAR WEIGHT
PEPTIDES
SULFUR 35
TRACER TECHNIQUES
AMINO ACIDS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
CARBOHYDRATES
CARBOXYLIC ACIDS
DAYS LIVING RADIOISOTOPES
DRUGS
EVEN-ODD NUCLEI
ISOTOPE APPLICATIONS
ISOTOPES
LIGHT NUCLEI
LIPOTROPIC FACTORS
NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC SULFUR COMPOUNDS
PROTEINS
RADIOISOTOPES
SACCHARIDES
SULFUR ISOTOPES
SYNTHESIS
550201* - Biochemistry- Tracer Techniques