The techniques of somatic cell genetics have been used to establish the linkage relationships of loci coding for two forms (A and B) of hexosaminidase (EC 3.2.1.30; 2-acetamido-2-deoxy-B-D-glucoside acetamidodeoxyglucohydrolase) and to determine whether a structural relationship exists between these forms. In a series of human-mouse hybrid cell lines, hexosaminidase A and B segregated independently. Our results and those reported by other investigators are used to analyze the proposed structural models for hexosaminidase. We have also been able to establish a syntenic relationship between the gene locus responsible for the expression of hexosaminidase A and those responsible for mannosephosphate isomerase and pyruvate kinase-3 and to assign the gene for hexosaminidase B to chromosome 5 in man. Tbere is thus a linkage between specific human autosomes and enzymes implicated in the production of lipid storage diseases.The lipid storage diseases are a family of inherited disorders characterized by the excessive accumulation of sphingolipids in the body's tissues. In each, the metabolic derangement appears to be the result of a deficiency of a specific lysosomal hydrolase which is involved in the catabolism of these complex lipids (1). One of these enzymes, P-N-acetylglucosaminidase (Hex; EC 3.2.1.30) is thought to be responsible for at least two lipodystrophies, Tay-Sachs' disease (TSD; GM2 gangliosidosis, type I) and Sandhoff's disease (SD; GM2 gangliosidosis, type II). When examined electrophoretically, this enzyme is found to exist in multiple forms, two of which (Hex A and B) have been well characterized biochemically (2). A third form of the enzyme (Hex C), about which relatively little is known, has recently been described (3). TSD is associated with a deficiency of Hex A and an increased activity of Hex B, and SD is associated with a deficiency of both Hex A and B (4,5). No individual has yet been reported in whom Hex A is present in the absence of Hex B.Biochemical, genetic, and immunological evidence suggests that a structural relationship exists between Hex A and B. Two theories concerning this relationship have recently been advanced (2, 6). The first proposes that Hex A is a conversion product of Hex B (2). TSD would then result from the deficiency of a functional conversion enzyme, and SD would re- sult from a defect in the gene coding for the basic Hex protein.The second theory proposes that Hex A and B are each composed of multiple subunits, one of which is common to both forms (6). In this hypothesis, TSD would result from the deficiency of the Hex A-specific subunit and SD from the deficiency of the common subunit. It is also possible that the two forms of Hex are not structurally related. Hex A and B may be controlled by two independent genes. TSD would then result from an effective deficiency of the normal Hex A structural gene product and SD might result from a mutation in a locus controlling expression of both enzymes or required for their activation. A series of human-mouse hybrid cell ...

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(19,20) suggested that there was no structural relationship between these two isozymes. We have therefore considered the possibility that the "Hex A" that occurs in mouse X human hybrids in the absence of human Hex B is, in point of fact, not human Hex A. Accordingly, we have investigated the "Hex A" activity that is found in the absence of Hex B in a human X mouse hybrid cell line that contains human chromosome 15 but not human chromosome 5. Immunological studies indicate that this "Hex A" material contains a normal human a subunit and is most probably a hybrid molecule containing human and mouse components. The genetic control of Tay-Sachs and Sandhoff diseases is discussed in the light of our findings. (23) and the genes for mannosephosphate isomerase, pyruvate kinase, and "Hex A" have been assigned to chromosome 15 (16-19). Five of the resulting subclones were analyzed for the continued expression of the human enzyme markers.Electrophoresis. Cells for Hex electrophoresis were grown in McCoy's 5A medium with 10% fetal calf serum added. They were harvested with 0.25% trypsin, centrifuged down, and resuspended in a 0.02 M citrate buffer, pH 5.2. The cell suspension was frozen and thawed five times and centrifuged at 14,500 X g for 15 min. The supernatant was concentrated to dryness by lyophilization. The final samples were dissolved in small volumes of 0.02 M citrate buffer, pH 5.2 (24). Cells for mannosephosphate isomerase, pyruvate kinase, and glucose-6-phosphate dehydrogenase electrophoresis were prepared as reported before (18).Starch gel electrophoresis was used to identify the human isozymes of mannosephosphate isomerase, pyruvate kinase, and glucose-6-phosphate dehydrogenase in the hybrid cell samples as described (18). Cellulose acetate gel [Cellogel; (17 cm X 17 cm X 0.5 mm) Kalex Scientific Co., Manhasset, N.Y.] was used to identify Hex A and B using modifications of the method of Okada and O'Brien (11). The sample solution (approximately 0.4 Al) was applied to the Cellogel with the aid of a multiple sample applicator (Shandon Scientific Co., Inc., Sewickley, Pa.). Electrophoresis was performed at 250 V for 2 hr, and the gel was incubated with 4-methylumbelliferyl-f.-D-glucosami iide (Sigma Chemical Co., St. Louis, Mo.), 0.2 mg/ml of 0.5 M citrate-phosphate buffer, pH 4.5. Hex activity was visualized as fluorescence under long wave UV light. Column Chromatography. Ion exchange chroma-. tography of Hex was carried out as described by Beutler et al. (8), except that the maximal NaCI concentration in the gradient was increased from 0.4 to 0.6 M.Immunological Studies. Rabbit antisera against human Hex

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Humans normally have 46 chromosomes in each cell, divided into 23 pairs. Two copies of chromosome 5, one copy inherited from each parent, form one of the pairs. Chromosome 5 spans about 181 million DNA building blocks (base pairs) and represents almost 6 percent of the total DNA in cells.