PMID- 10767175
OWN - NLM
STAT- MEDLINE
DCOM- 20000531
LR  - 20131121
IS  - 1096-7192 (Print)
IS  - 1096-7192 (Linking)
VI  - 69
IP  - 3
DP  - 2000 Mar
TI  - The V388M mutation results in a kinetic variant form of phenylalanine 
      hydroxylase.
PG  - 204-12
AB  - The molecular mechanism underlying the metabolic defect in phenylketonuria (PKU) 
      patients carrying the V388M missense mutation of the phenylalanine hydroxylase 
      (PAH) gene has been characterized. An in vitro prokaryotic expression system has 
      been used to produce both the wild-type and the mutant form of the human PAH 
      (hPAH) protein. The recombinant enzymes, obtained as fusion proteins, were 
      purified by immobilized metal affinity chromatography and recovered in high 
      yields. The wild-type hPAH possessed a high specific activity and its kinetic 
      properties were the same as those reported for the enzyme isolated from human 
      liver and other recombinant wild-type hPAH enzymes. The recombinant V388M mutant 
      form exhibited a reduced specific activity equivalent to 30% of the wild-type 
      hPAH enzyme when assayed using the synthetic cofactor (6-methyltetrahydropterin). 
      Lower values were obtained (23 and 19%) when the mutant enzyme was assayed with 
      the natural cofactor ((6R)-tetrahydrobiopterin) and different concentrations of 
      l-phenylalanine. The enzyme kinetic studies of the V388M mutant protein revealed 
      that this enzyme was a kinetic variant form of hPAH with a reduced affinity for 
      l-phenylalanine and for the natural cofactor ((6R)-tetrahydrobiopterin). The 
      residual activities determined for the V388M form of hPAH were compatible with 
      the phenotype presented by the PKU patients harboring the V388M mutation in the 
      PAH gene.
CI  - Copyright 2000 Academic Press.
FAU - Leandro, P
AU  - Leandro P
AD  - Centro de Patogenese Molecular, Faculdade de Farmacia, University of Lisboa, 
      Lisboa, 1600, Portugal.
FAU - Rivera, I
AU  - Rivera I
FAU - Lechner, M C
AU  - Lechner MC
FAU - de Almeida, I T
AU  - de Almeida IT
FAU - Konecki, D
AU  - Konecki D
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - United States
TA  - Mol Genet Metab
JT  - Molecular genetics and metabolism
JID - 9805456
RN  - 0 (Recombinant Fusion Proteins)
RN  - 47E5O17Y3R (Phenylalanine)
RN  - EC 1.14.16.1 (Phenylalanine Hydroxylase)
SB  - IM
MH  - Amino Acid Substitution
MH  - Gene Expression Regulation, Enzymologic
MH  - Genetic Variation
MH  - Humans
MH  - Kinetics
MH  - Mutation
MH  - Phenylalanine/metabolism
MH  - Phenylalanine Hydroxylase/*genetics/isolation & purification/metabolism
MH  - Recombinant Fusion Proteins/genetics/isolation & purification/metabolism
EDAT- 2000/04/18 09:00
MHDA- 2000/06/03 09:00
CRDT- 2000/04/18 09:00
PHST- 2000/04/18 09:00 [pubmed]
PHST- 2000/06/03 09:00 [medline]
PHST- 2000/04/18 09:00 [entrez]
AID - S1096-7192(00)92970-6 [pii]
AID - 10.1006/mgme.2000.2970 [doi]
PST - ppublish
SO  - Mol Genet Metab. 2000 Mar;69(3):204-12. doi: 10.1006/mgme.2000.2970.