IDENTIFICATION OF AMINO-ACID CHANGES AFFECTING YEAST UROPORPHYRINOGEN DECARBOXYLASE ACTIVITY BY SEQUENCE-ANALYSIS OF HEM12 MUTANT ALLELES

被引:11
作者
CHELSTOWSKA, A
ZOLADEK, T
GAREY, J
KUSHNER, J
RYTKA, J
LABBEBOIS, R
机构
[1] UNIV PARIS 07, INST JACQUES MONOD, BIOCHIM PORPHYRINES LAB, 2 PL JUSSIEU, F-75251 PARIS 05, FRANCE
[2] POLISH ACAD SCI, INST BIOCHEM & BIOPHYS, PL-02532 WARSAW, POLAND
[3] UNIV UTAH, SCH MED, DIV HEMATOL & ONCOL, SALT LAKE CITY, UT 84132 USA
关键词
D O I
10.1042/bj2880753
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The molecular basis of the uroporphyrinogen decarboxylase defect in eleven yeast 'uroporphyric' mutants was investigated. Uroporphyrinogen decarboxylase, an enzyme of the haem-biosynthetic pathway, catalyses the decarboxylation of uroporphyrinogen to coproporphyrinogen and is encoded by the HEM12 gene in the yeast Saccharomyces cerevisiae. The mutations were identified by sequencing the mutant hem12 alleles amplified in vitro from genomic DNA extracted from the mutant strains. Four mutations leading to the absence of enzyme protein were found: one mutation caused the substitution of the translation initiator Met to Ile, a two-base deletion created a frameshift at codon 247 and two nonsense mutations were found at codons 50 and 263. Four different point mutations were identified in seven 'leaky' mutants with residual modified uroporphyrinogen decarboxylase activity; each of three mutations was found in two independently isolated mutants. The nucleotide transitions resulted in the amino acid substitutions Ser-59 to Phe, Thr-62 to Ile, Leu-107 to Ser, or Ser-215 to Asn, all located in or near highly conserved regions. The results suggest that there is a single active centre in uroporphyrinogen decarboxylase, the geometry of which is affected in the mutant enzymes.
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页码:753 / 757
页数:5
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