Manoj Kelath Murali, Parashar Abhinav, Venkatachalam Avanthika, Goyal Sahil, Singh Preeti Gunjan, Gade Sudeep K, Periyasami Kalaiselvi, Jacob Reeba Susan, Sardar Debosmita, Singh Shanikant, Kumar Rajan, Gideon Daniel A
Satyamjayatu: The Science & Ethics Foundation, Kulappully, Shoranur-2, Kerala 679122, India; Hemoproteins Lab, School of Bio Sciences and Technology, VIT University, Vellore, Tamil Nadu 632014, India; REDOx Lab, PSG Institute of Advanced Studies, Coimbatore, Tamil Nadu 641004, India; School of Chemical and Bio Technology, SASTRA University, Thanjavur, Tamil Nadu 613401, India; Late Lowell P. Hager's Lab, Department of Biochemistry, University of Illinois (U-C), 600, S Mathews Avenue, Urbana, IL 61801, USA.
Hemoproteins Lab, School of Bio Sciences and Technology, VIT University, Vellore, Tamil Nadu 632014, India.
Biochimie. 2016 Jun;125:91-111. doi: 10.1016/j.biochi.2016.03.003. Epub 2016 Mar 10.
Peroxidations mediated by heme-enzymes have been traditionally studied under a single-site (heme distal pocket), non-sequential (ping-pong), two-substrates binding scheme of Michaelis-Menten paradigm. We had reported unusual modulations of peroxidase and P450 reaction outcomes and explained it invoking diffusible reactive species [Manoj, 2006; Manoj et al., 2010; Andrew et al., 2011, Parashar et al., 2014 & Venkatachalam et al., 2016].
A systematic investigation of specific product formation rates was undertaken to probe the hypothesis that involvement of diffusible reactive species could explain undefined substrate specificities and maverick modulations (sponsored by additives) of heme-enzymes.
When the rate of specific product formation was studied as a function of reactants' concentration or environmental conditions, we noted marked deviations from normal profiles. We report that heme-enzyme mediated peroxidations of various substrates are inhibited (or activated) by sub-equivalent concentrations of diverse redox-active additives and this is owing to multiple redox equilibriums in the milieu. At low enzyme and peroxide concentrations, the enzyme is seen to recycle via a one-electron (oxidase) cycle, which does not require the substrate to access the heme centre. Schemes are provided that explain the complex mechanistic cycle, kinetics & stoichiometry.
It is not obligatory for an inhibitor or substrate to interact with the heme centre for influencing overall catalysis. Roles of diffusible reactive species explain catalytic outcomes at low enzyme and reactant concentrations.
The current work highlights the scope/importance of redox enzyme reactions that could occur "out of the active site" in biological or in situ systems.
传统上，血红素酶介导的过氧化反应是在米氏范式的单位点（血红素远端口袋）、非顺序（乒乓）、双底物结合模式下进行研究的。我们曾报道过过氧化物酶和细胞色素P450反应结果的异常调节，并通过可扩散反应物种对此进行了解释[马诺伊，2006年；马诺伊等人，2010年；安德鲁等人，2011年，帕拉沙尔等人，2014年及文卡塔查拉姆等人，2016年]。
对特定产物形成速率进行了系统研究，以探究可扩散反应物种的参与能否解释血红素酶未明确的底物特异性和异常调节（由添加剂引发）这一假设。
当将特定产物形成速率作为反应物浓度或环境条件的函数进行研究时，我们注意到与正常曲线存在明显偏差。我们报告称，各种底物的血红素酶介导的过氧化反应会受到亚等效浓度的多种氧化还原活性添加剂的抑制（或激活），这是由于环境中的多种氧化还原平衡所致。在低酶和过氧化物浓度下，酶通过单电子（氧化酶）循环进行循环利用，该循环不需要底物进入血红素中心。提供了能够解释复杂机理循环、动力学和化学计量的示意图。
抑制剂或底物并非必须与血红素中心相互作用才能影响整体催化作用。可扩散反应物种的作用解释了低酶和反应物浓度下的催化结果。
当前工作突出了生物或原位系统中可能在“活性位点之外”发生的氧化还原酶反应的范围/重要性。
