Hydrophobic binding domain of rat intestinal maltase-glucoamylase
Conference
·
· Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)
OSTI ID:5127257
Maltase-glucoamylase is an amphiphilic oligomeric ectoenzyme on the intestinal brush border. The enzyme was isolated by Triton X-100 extraction, purified in the presence of protease inhibitors and incorporated into phospholipid vesicles. After purification of the vesicles on Sephadex G-50, maltase was labelled with a photoactive reagent (/sup 125/I TID). The labelled enzyme was extracted with acetone, reincorporated into phospholipid vesicles and digested with activated papain. Vesicle bound and free enzyme were separated on Sepharose 4B. 90% of the enzyme activity was free, while 90% of the /sup 125/I label remained vesicle bound, indicating separation of an active polar headpiece from a labelled apolar peptide in the lipid bilayer. The vesicle fractions were chromatographed on Sephadex LH-60 with ethanol-formic acid (7:3, v/v). A single radioactive peak (Mr 14 kDa) was separated from labelled lipid. SDS-PAGE of the peak showed a radioactive doublet of 26-28 kDa, possibly representing a dimer. No other labelled peptides were found. These results suggest that detergent-solubilized maltase-glucoamylase is inserted into the phospholipid bilayer via an apolar peptide with a minimum molecular weight of 14 kDa. The peptide is probably on the 145 kDa subunit which is converted to 130 kDa when the membrane bound enzyme is solubilized by papain.
- Research Organization:
- Univ. of Toronto, Ontario
- OSTI ID:
- 5127257
- Report Number(s):
- CONF-8606151-
- Conference Information:
- Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States) Journal Volume: 45:6
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMYLASE
ANIMALS
BETA DECAY RADIOISOTOPES
BODY
CHROMATOGRAPHY
DAYS LIVING RADIOISOTOPES
DIGESTIVE SYSTEM
ELECTRON CAPTURE RADIOISOTOPES
ENZYME ACTIVITY
ENZYMES
GASTROINTESTINAL TRACT
GLYCOSYL HYDROLASES
HYDROLASES
INTERMEDIATE MASS NUCLEI
INTESTINES
IODINE 125
IODINE ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
LABELLING
MAMMALS
MOLECULAR WEIGHT
NUCLEI
O-GLYCOSYL HYDROLASES
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
ORGANS
PEPTIDES
PROTEINS
PURIFICATION
RADIOISOTOPES
RATS
RODENTS
SEPARATION PROCESSES
TRACER TECHNIQUES
VERTEBRATES
59 BASIC BIOLOGICAL SCIENCES
AMYLASE
ANIMALS
BETA DECAY RADIOISOTOPES
BODY
CHROMATOGRAPHY
DAYS LIVING RADIOISOTOPES
DIGESTIVE SYSTEM
ELECTRON CAPTURE RADIOISOTOPES
ENZYME ACTIVITY
ENZYMES
GASTROINTESTINAL TRACT
GLYCOSYL HYDROLASES
HYDROLASES
INTERMEDIATE MASS NUCLEI
INTESTINES
IODINE 125
IODINE ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
LABELLING
MAMMALS
MOLECULAR WEIGHT
NUCLEI
O-GLYCOSYL HYDROLASES
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
ORGANS
PEPTIDES
PROTEINS
PURIFICATION
RADIOISOTOPES
RATS
RODENTS
SEPARATION PROCESSES
TRACER TECHNIQUES
VERTEBRATES