Two forms of the lysosomal enzyme beta-mannosidase were identified and purified from human urine. The purification strategy employed allowed sufficient quantities of both forms to be obtained for subunit analysis and for further characterizations. The two beta-mannosidases were identified as beta-mannosidase B and A, in order of their elution from an ion-exchange column. In all samples examined, the A form was predominant, and the B/A ratio was consistently 0.14. The two forms displayed the same optimum pH (i.e., 4.3) and both were retained by a Concanavalin-A Sepharose column, but showed different isoelectric points, molecular masses and subunit compositions. Native- and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analyses of pure beta-mannosidases B and A suggest that active protein B (160 kDa) consists of three subunits, one 75 kDa and two 49 kDa subunits. Protein A is smaller and appears to be composed of three subunits of 75 kDa, 49 kDa and 37 kDa. Two forms of beta-mannosidase, exhibiting a chromatographic behaviour comparable to the urinary forms, were also detected in human kidney. Nevertheless, in this tissue their relative distribution was different, the B/A ratio being 19.

Guadalupi, R., Bernard, M., Orlacchio, A., Foglietti, M., & Emiliani, C. (1996). Purification and properties of human urinary beta-D-mannosidase. BIOCHIMICA ET BIOPHYSICA ACTA-PROTEIN STRUCTURE AND MOLECULAR ENZYMOLOGY, 1293(1), 9-16 [10.1016/0167-4838(95)00225-1].

Purification and properties of human urinary beta-D-mannosidase

ORLACCHIO, ANTONIO;
1996-03-07

Abstract

Two forms of the lysosomal enzyme beta-mannosidase were identified and purified from human urine. The purification strategy employed allowed sufficient quantities of both forms to be obtained for subunit analysis and for further characterizations. The two beta-mannosidases were identified as beta-mannosidase B and A, in order of their elution from an ion-exchange column. In all samples examined, the A form was predominant, and the B/A ratio was consistently 0.14. The two forms displayed the same optimum pH (i.e., 4.3) and both were retained by a Concanavalin-A Sepharose column, but showed different isoelectric points, molecular masses and subunit compositions. Native- and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analyses of pure beta-mannosidases B and A suggest that active protein B (160 kDa) consists of three subunits, one 75 kDa and two 49 kDa subunits. Protein A is smaller and appears to be composed of three subunits of 75 kDa, 49 kDa and 37 kDa. Two forms of beta-mannosidase, exhibiting a chromatographic behaviour comparable to the urinary forms, were also detected in human kidney. Nevertheless, in this tissue their relative distribution was different, the B/A ratio being 19.
Pubblicato
Rilevanza internazionale
Articolo
Esperti anonimi
Settore BIO/12
English
Con Impact Factor ISI
Mannosidases; Electrophoresis, Polyacrylamide Gel; Humans; Hydrogen-Ion Concentration; Isoelectric Point; Temperature; Protein Denaturation; Molecular Weight; beta-Mannosidase; Chromatography, DEAE-Cellulose; Enzyme Stability; Chromatography, Agarose; Isoenzymes; Kidney; Lysosomes; Protein Conformation
Guadalupi, R., Bernard, M., Orlacchio, A., Foglietti, M., & Emiliani, C. (1996). Purification and properties of human urinary beta-D-mannosidase. BIOCHIMICA ET BIOPHYSICA ACTA-PROTEIN STRUCTURE AND MOLECULAR ENZYMOLOGY, 1293(1), 9-16 [10.1016/0167-4838(95)00225-1].
Guadalupi, R; Bernard, M; Orlacchio, A; Foglietti, M; Emiliani, C
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/2108/90070
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