caa a22 4500 445821221 CHVBK 20180317145244.0 cr unu---uuuuu 170323e20110301xx s 000 0 eng 10.1007/s00239-011-9433-8 doi (NATIONALLICENCE)springer-10.1007/s00239-011-9433-8 Evolution of Metamorphism in Thymidylate Synthases Within the Primate Lineages [Elektronische Daten] [BeiBei Luo, Saphronia Johnson, Lukasz Lebioda, Sondra Berger] Crystal structures of human thymidylate synthase (hTS) revealed that the protein exists in active and inactive conformations, defined by the position of a loop containing the active site nucleophile. TS is highly homologous among diverse species; however, the residue at position 163 (hTS) differs among species. Arginine at this position is predicted by structural modeling to enable conformational switching. Arginine or lysine is reported at this position in all mammals in the GenBank and Ensembl databases, with arginine reported in only primates. Sequence analysis of the TS gene of representative primates revealed that arginine occurs at this relative position in all primates except a representative of prosimians. Mutant human proteins were created with residues at position 163 that occur in TSs from prokaryotes and eukaryotes. Catalytic constants (k cat) of mutant enzymes were 45-149% of hTS, with the lysine mutant (R163K) exhibiting the highest k cat. The effect of lysine substitution on solution structure and on ligand binding was investigated. R163K exhibited higher intrinsic fluorescence, a more negative molar ellipticity, and higher dissociation constants (K d) for ligands that modulate protein conformation than hTS. Temperature effects on intrinsic fluorescence and catalytic activity of hTS and R163K are consistent with proteins populating different conformational states. The data indicate that the enzyme with arginine at the position corresponding to 163 (hTS) evolved after the divergence of prosimians and simians and that substitution of lysine by arginine confers unique structural and functional properties to the enzyme expressed in simian primates. Springer Science+Business Media, LLC, 2011 Protein metamorphism nationallicence Enzyme catalysis nationallicence Primate evolution nationallicence Intrinsic fluorescence nationallicence Conformational switching nationallicence Protein structure nationallicence Energy of activation nationallicence TS : Thymidylate synthase (h, human nationallicence ec, Escherichia coli) nationallicence mTHF : 5,10-Methylenetetrahydrofolate nationallicence DHF : Dihydrofolate nationallicence ZD1694 : N-(5-[N-(3,4-Dihydro-2-methyl-4-oxyquinazolin-6-ylmethyl)-N-methyl-amino]-2-thenoyl)-l-glutamic acid (raltitrexed, Tomudex) nationallicence CD : Circular dichroism nationallicence Luo BeiBei Department of Pharmaceutical and Biomedical Sciences, South Carolina College of Pharmacy, University of South Carolina, 700 Sumter Street, 29208, Columbia, SC, USA aut Johnson Saphronia Department of Pharmaceutical and Biomedical Sciences, South Carolina College of Pharmacy, University of South Carolina, 700 Sumter Street, 29208, Columbia, SC, USA aut Lebioda Lukasz Department of Chemistry and Biochemistry, University of South Carolina, 29208, Columbia, SC, USA aut Berger Sondra Department of Pharmaceutical and Biomedical Sciences, South Carolina College of Pharmacy, University of South Carolina, 700 Sumter Street, 29208, Columbia, SC, USA aut Journal of Molecular Evolution Springer-Verlag 72/3(2011-03-01), 306-314 0022-2844 72:3<306 2011 72 239 https://doi.org/10.1007/s00239-011-9433-8 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s00239-011-9433-8 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Luo BeiBei Department of Pharmaceutical and Biomedical Sciences, South Carolina College of Pharmacy, University of South Carolina, 700 Sumter Street, 29208, Columbia, SC, USA aut NATIONALLICENCE 700 1- Johnson Saphronia Department of Pharmaceutical and Biomedical Sciences, South Carolina College of Pharmacy, University of South Carolina, 700 Sumter Street, 29208, Columbia, SC, USA aut NATIONALLICENCE 700 1- Lebioda Lukasz Department of Chemistry and Biochemistry, University of South Carolina, 29208, Columbia, SC, USA aut NATIONALLICENCE 700 1- Berger Sondra Department of Pharmaceutical and Biomedical Sciences, South Carolina College of Pharmacy, University of South Carolina, 700 Sumter Street, 29208, Columbia, SC, USA aut NATIONALLICENCE 773 0- Journal of Molecular Evolution Springer-Verlag 72/3(2011-03-01), 306-314 0022-2844 72:3<306 2011 72 239 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer