Bovine kidney beta-mannosidase: purification and characterization

Biochem J. 1993 Jan 15;289 ( Pt 2)(Pt 2):343-7. doi: 10.1042/bj2890343.

Abstract

Lysosomal beta-mannosidase was purified 160,000-fold in 24% yield from bovine kidney by a four-step purification procedure, which included concanavalin A-Sepharose, immunoaffinity, TSK-butyl and h.p.l.c. cation-exchange chromatography. When analysed by SDS/PAGE and detected by Coomassie Blue or silver staining, the purified enzyme preparation consists of two prominent peptides (100 and 110 kDa) and a third minor peptide (84 kDa). These three peptides are immunologically related and are consistently associated with beta-mannosidase activity in all chromatographic steps. Removal of N-linked carbohydrate from the 84, 100 and 110 kDa peptides decreases their molecular sizes to 75, 86 and 91 kDa respectively. Bovine kidneys lacking beta-mannosidase, activity, acquired from calves affected with beta-mannosidosis, do not contain detectable quantities of the three beta-mannosidase peptides, as judged by monoclonal- and polyclonal-antibody reactivity.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Carbohydrates / analysis
  • Cattle
  • Cattle Diseases
  • Chromatography, Affinity
  • Chromatography, High Pressure Liquid
  • Chromatography, Ion Exchange
  • Electrophoresis, Polyacrylamide Gel
  • Kidney / enzymology*
  • Kinetics
  • Lysosomes / enzymology
  • Mannosidases / genetics
  • Mannosidases / isolation & purification*
  • Mannosidases / metabolism
  • Molecular Weight
  • Reference Values
  • alpha-Mannosidosis / enzymology
  • alpha-Mannosidosis / genetics
  • alpha-Mannosidosis / veterinary
  • beta-Mannosidase

Substances

  • Carbohydrates
  • Mannosidases
  • beta-Mannosidase