Characterization of intracellular pteroylpolyglutamate hydrolase (PPH) from human intestinal mucosa
There are two forms of pteroylpolyglutamate hydrolase (PPH) in the human intestinal mucosa, one in the brush border membrane and the other intracellular; brush border PPH is an exopeptidase with optimal activity at pH 6.5 and a requirement for zinc. The presence study characterized human intracellular PPH and compared its properties to those of brush border PPH. Intracellular PPH was purified 30-fold. The enzyme had a MW of 75,000 by gel filtration, was optimally active at pH 4.5, and had an isoelectric point at pH 8.0. In contrast to brush border PPH, intracellular PPH was unstable at increasing temperatures, was unaffected by dialysis against chelating agents and showed no requirement for Zn/sup 2 +/. Using PteGlu/sub 2/(/sup 14/C)Glu as substrate, they demonstrated a K/sub m/ of 1.2 ..mu..M and increasing affinity for folates with longer glutamate chains. Intracellular PPH required the complete folic acid (PteGlu) moiety and a ..gamma..-glutamyl linkage for activity. Using ion exchange chromatography and an HPLC method to determine the hydrolytic products of the reaction, they found intracellular PPH could cleave both internal and terminal ..gamma..-glutamyl linkages, with PteGlu as an end product. After subcellular fractionation of the mucosa, PPH was found in the lysosomes. In summary, the distinct characteristics of brush border and intracellular PPH suggest that the two hydrolases serve different roles in folate metabolism.
- Research Organization:
- Univ. of California, Davis
- OSTI ID:
- 5462710
- Report Number(s):
- CONF-8604222-; TRN: 86-028381
- Journal Information:
- Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Vol. 45:3; Conference: 70. annual meeting of the Federation of American Society for Experimental Biology, St. Louis, MO, USA, 13 Apr 1986
- Country of Publication:
- United States
- Language:
- English
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FOLIC ACID
METABOLISM
HYDROLASES
ENZYME ACTIVITY
AFFINITY
CARBON 14 COMPOUNDS
CHELATING AGENTS
COMPARATIVE EVALUATIONS
INTESTINES
ION EXCHANGE CHROMATOGRAPHY
LIQUID COLUMN CHROMATOGRAPHY
LYSOSOMES
MAN
MUCOUS MEMBRANES
TEMPERATURE DEPENDENCE
TRACER TECHNIQUES
ZINC
AMINO ACIDS
ANIMALS
AROMATICS
AZAARENES
BODY
CARBOXYLIC ACIDS
CELL CONSTITUENTS
CHROMATOGRAPHY
DIGESTIVE SYSTEM
DRUGS
ELEMENTS
ENZYMES
GASTROINTESTINAL TRACT
HEMATINICS
HEMATOLOGIC AGENTS
HETEROCYCLIC COMPOUNDS
HYDROXY COMPOUNDS
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
MAMMALS
MEMBRANES
METALS
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORGANOIDS
ORGANS
PRIMATES
PTERIDINES
SEPARATION PROCESSES
VERTEBRATES
VITAMIN B GROUP
VITAMINS
550201* - Biochemistry- Tracer Techniques