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2002 | 49 | 3 | 651-658
Article title

The effect of Arg209 to Lys mutation in mouse thymidylate synthase.

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EN
Abstracts
EN
Mouse thymidylate synthase R209K (a mutation corresponding to R218K in Lactobacillus casei), overexpressed in thymidylate synthase-deficient Escherichia coli strain, was poorly soluble and with only feeble enzyme activity. The mutated protein, incubated with FdUMP and N5,10-methylenetetrahydrofolate, did not form a complex stable under conditions of SDS/polyacrylamide gel electrophoresis. The reaction catalyzed by the R209K enzyme (studied in a crude extract), compared to that catalyzed by purified wild-type recombinant mouse thymidylate synthase, showed the Km value for dUMP 571-fold higher and Vmax value over 50-fold (assuming that the mutated enzyme constituted 20% of total crude extract protein) lower. Thus the ratios kcat, R209K/kcat, 'wild' and (kcat, R209K/Km, R209KdUMP)/( kcat, 'wild'/Km, 'wild'dUMP) were 0.019 and 0.000032, respectively, documenting that mouse thymidylate synthase R209, similar to the corresponding L. casei R218, is essential for both dUMP binding and enzyme reaction.
Publisher

Year
Volume
49
Issue
3
Pages
651-658
Physical description
Dates
published
2002
received
2002-07-23
accepted
2002-08-27
Contributors
  • >Nencki Institute of Experimental Biology, Warszawa, Poland
  • >Nencki Institute of Experimental Biology, Warszawa, Poland
  • Institute of Biology and Environmental Protection, Rzeszów University, Rzeszów, Poland
  • >Nencki Institute of Experimental Biology, Warszawa, Poland
  • Institute of Biotechnology and Antibiotics, Warszawa, Poland
author
  • >Nencki Institute of Experimental Biology, Warszawa, Poland
References
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Document Type
Publication order reference
Identifiers
YADDA identifier
bwmeta1.element.bwnjournal-article-abpv49i3p651kz
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