Beta-mannosidase, a lysosomal enzyme which acts exclusively at the last step of oligosaccharide catabolism in glycoprotein degradation, functions to cleave the single beta-linked mannose sugar found in all N-linked oligosaccharides of glycoproteins. Deficiency of this enzyme results in beta-mannosidosis, a lysosomal storage disease characterized by the cellular accumulation of small oligosaccharides. This autosomal recessive genetic disease is the most recently identified human acid glycosyl hydrolase deficiency. In the human beta-mannosidosis cases described to date the clinical presentation is relatively mild for a lysosomal storage disease and very variable. Nothing is known about the molecular basis for human beta-mannosidosis. The reasons for the variability are unknown and will be addressed in this proposal. Bovine beta-mannosidase cDNA has been cloned and sequenced. A full length cDNA clone has been prepared and verified by its in vitro expression. Using this information, it is now possible to isolate and characterize the normal human beta-mannosidase cDNA and gene, to identify the mutations that cause beta-mannosidosis, and to explore the molecular pathology of this disease.
The specific aims of this proposal ar: 1. Clone and characterize cDNA for human beta-mannosidase. Human beta-mannosidase cDNA will be cloned using information obtained from the bovine cDNA. A full length human cDNA will be constructed for expression in transfected cells. 2. Clone and characterize the human genomic beta-mannosidase sequence. The genomic structure of the beta- mannosidase gene from any species is not yet known. We plan to clone the human beta-mannosidase gene, determine its transcriptional start site(s) and intron/exon placement in the gene. Achievement of specific aims 1 and 2 will characterized and will provide the reagents required to peruse mutation analysis and structure-function analysis. 3. Determine the molecular pathology to human beta-mannosidosis. The levels of beta-mannosidase protein and mRNA will be evaluated in fibroblast cell cultures derived from beta-mannosidase patients. The mutations responsible for each case of beta-mannosidosis will be identified and correlated with the severity and symptoms in the presentation of the disease. Experiments in Specific aim 3 will delineate the mutations which cause the human disease to provide insight into the nature of the extreme variability of the human disease. Information of the molecular pathology of beta-mannosidosis will allow prediction about the clinical spectrum of the disease and suggest where to look for additional cases of this potentially underreported disease. Structure-function studies will lead to further understanding of the enzyme providing information useful for potential treatment of diseases involving glycosyl hydrolases.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
First Independent Research Support & Transition (FIRST) Awards (R29)
Project #
5R29DK049782-02
Application #
2518479
Study Section
Medical Biochemistry Study Section (MEDB)
Program Officer
Mckeon, Catherine T
Project Start
1996-09-15
Project End
2001-08-31
Budget Start
1997-09-01
Budget End
1998-08-31
Support Year
2
Fiscal Year
1997
Total Cost
Indirect Cost
Name
Michigan State University
Department
Pathology
Type
Schools of Medicine
DUNS #
193247145
City
East Lansing
State
MI
Country
United States
Zip Code
48824