Substrate concentration dependence of deuterium isotope effects on beef heart lactate dehydrogenase |
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| Authors: | A M Cantwell D Dennis |
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| Affiliation: | 1. Laboratory for Orthopaedic Pathophysiology and Regenerative Medicine, IRCCS Istituto Ortopedico Rizzoli, Via di Barbiano 1/10, 40136 Bologna, Italy;2. Department of Biomedical and Neuromotor Sciences, University of Bologna, Italy;3. Department of Applied Science and Technology, Politecnico di Torino, Corso Duca degli Abruzzi 24, Torino 10129, Italy;4. Department of Clinical and Molecular Sciences (DISCLIMO), Università Politecnica delle Marche, Via Tronto 10/a, Ancona 60020, Italy;1. School of Biochemistry and Immunology, Trinity College Dublin, Dublin, Ireland;2. School of Medicine, Trinity College Dublin, Dublin, Ireland;3. Trinity Centre for Bioengineering, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland;4. Department of Mechanical and Manufacturing Engineering, School of Engineering, Trinity College Dublin, Dublin, Ireland;5. Advanced Materials and Bioengineering Research (AMBER) Centre, Trinity College Dublin, Ireland |
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| Abstract: | The kinetic effect of substituting deuterium for the transferred hydrogen at the C-2 position of L-lactic acid was studied by measuring the initial velocity (vi) of lactate oxidation by beef-heart lactate dehydrogenase (L-lactate: NAD oxidoreductase, EC 1.1.1.27).viH/viD values were found to decrease with increasing substrate concentration. At saturating levels of substrate VMH/VMD was equal to one. This is an example of the masking of an isotope effect by a slow step in the reaction sequence which is not affected by deuterium substitution. Such a result is evidence that the covalent transfer of hydrogen is not the rate-limiting step in lactate oxidation. Measurement of viH/viD at low substrate concentration gives an estimate of the importance of steps known to be affected by deuterium substitution in the overall rate expression. |
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