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One amino acid makes a difference-Characterization of a new TPMT allele and the influence of SAM on TPMT stability.

ABSTRACT: Thiopurine induced toxicity is associated with defects in the thiopurine methyltransferase (TPMT) gene. TPMT is a polymorphic enzyme, with most of the single nucleotide polymorphisms (SNPs) causing an amino acid change, altering the enzymatic activity of the TPMT protein. In this study, we characterize a novel patient allele c.719A?>?C, named TPMT*41, together with the more common variant *3C c.719A?>?G, resulting in an amino acid shift at tyrosine 240 to serine, p.Y240S and cysteine, p.Y240C respectively. We show that the patient heterozygote for c.719A?>?C has intermediate enzymatic activity in red blood cells. Furthermore, in vitro studies, using recombinant protein, show that TPMT p.Y240S is less stable than both TPMTwt and TPMT p.Y240C. The addition of SAM increases the stability and, in agreement with Isothermal Titration Calorimetry (ITC) data, higher molar excess of SAM is needed in order to stabilize TPMT p.Y240C and TPMT p.Y240S compared to TPMTwt. Molecular dynamics simulations show that the loss of interactions is most severe for Y240S, which agrees with the thermal stability of the mutations. In conclusion, our study shows that SAM increases the stability of TPMT and that changing only one amino acid can have a dramatic effect on TPMT stability and activity.

SUBMITTER: Iu YPH

PROVIDER: S-EPMC5411961 | biostudies-literature | 2017 May

REPOSITORIES: biostudies-literature

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One amino acid makes a difference-Characterization of a new TPMT allele and the influence of SAM on TPMT stability.

Iu Yan Ping Heidi YPH Helander Sara S Kahlin Anna Zimdahl AZ Cheng Chun Wah CW Shek Chi Chung CC Leung Moon Ho MH Wallner Björn B Mårtensson Lars-Göran LG Appell Malin Lindqvist ML

Scientific reports 20170502

Thiopurine induced toxicity is associated with defects in the thiopurine methyltransferase (TPMT) gene. TPMT is a polymorphic enzyme, with most of the single nucleotide polymorphisms (SNPs) causing an amino acid change, altering the enzymatic activity of the TPMT protein. In this study, we characterize a novel patient allele c.719A > C, named TPMT*41, together with the more common variant *3C c.719A > G, resulting in an amino acid shift at tyrosine 240 to serine, p.Y240S and cysteine, p.Y240C re ...[more]

PMID: 28462921

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Dataset Information

One amino acid makes a difference-Characterization of a new TPMT allele and the influence of SAM on TPMT stability.

Publications

One amino acid makes a difference-Characterization of a new TPMT allele and the influence of SAM on TPMT stability.

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