LEUKOCYTE BETA-GLUCOSIDASE IN HOMOZYGOTES AND HETEROZYGOTES FOR GAUCHER DISEASE

  • 1 January 1980
    • journal article
    • research article
    • Vol. 32  (2) , 158-173
Abstract
Human leukocytes contain at least 2 isozymes of 4-methylumbelliferyl-.beta.-glucosidase acting optimally at pH 4.0 and 4.8; in Gaucher disease, only the former is deficient. Brief exposure of the leukocyte homogenate to pH 4.0 at room temperature results in irreversible inactivation of the pH 4.8 activity, while the activity at pH 4.0 remains unaffected. The more acidic isozyme is stimulated 4- to 5-fold by 0.2% sodium taurodeoxycholate (TDC) with a shift in the pH optimum to 5.0. The less acidic isozyme is completely suppressed in the presence of this detergent. Both leukocyte isozymes appear to be membrane-bound since gel filtration of Sephadex G-200 produces only 1 peak of activity located at the void volume, unlike in liver and kidney where a 2nd peak can also be demonstrated. Heat inactivation analysis indicated that in controls, assayed in the absence of detergent, pH 4.0 activity is more thermostable than pH 4.8 activity. In Gaucher disease, the residual .beta.-glucosidase at pH 4.0 is just as thermolabile as the unaffected pH 4.8 activity. Heat inactivation of the enzyme in the presence of TDC resulted in rapid loss of activity, suggesting a direct effect of the bile salt on the configuration of the enzyme decreasing its thermal stability. In the absence of detergent, acid .beta.-glucosidase shows 2 Km, one at 3.2 mM and another at 0.9 mM. In the presence of detergent, only the higher Km at 3.3 mM is obtained. In patients with Gaucher disease and in obligate carriers, the Km remains essentially unaffected while the Vmax shows the expected deficiency. A reliable and reproducible selective assay technique was developed for the diagnosis of Gaucher disease homozygotes and obligate heterozygotes and for the carrier screening of individuals at risk for this inherited disorder. The efficacy of this technique was demonstrated by studying the activity in 42 controls, 26 patients, 32 obligate heterozygotes and 23 healthy relatives of patients with Gaucher disease.