Model complexes of active sites in carbonic anhydrase to facilitate the hydration of carbon dioxide to carbonate

박동국 국민대학교일반대학원 화학과 무기화학전공 2016 국내박사

A rapid global warming and climate change need global efforts to reduce the concentration of atmospheric carbon dioxide. Carbon capture and storage technology (CCS) can help mitigate CO2 emissions from large emitters such as steel plants, cement plants and thermoelectric power plants. However, CO2 is an inert and unreactive molecule that is thermodynamically stable, which means it difficult to capture and separate. The chemical absorption of CO2 using a monoethanolamine (amines) solvent is currently the most widely accepted commercial approach. However, desorption of CO2 for recovery of the solvent requires a large parasitic high supply of energy. Therefore, alternative process for CO2 absorption has been recently proposed to replace the amine solvents with an eco-friendly biocatalyst. The proposed eco-friendly biocatalyst mainly used carbonic anhydrase (CA). CA is a ubiquitous enzyme that catalyzes the conversion CO2 to bicarbonate or vice versa. The use of CA for CO2 capture can potentially overcome the limitations of amine solvents. In spite of unparalleled advantages, it is difficult to apply to commercial CO2 capture processes because of expensive production costs, short life time, temperature and pH sensitive and so on. In order to overcome such potential limitations of CA, this thesis study the small molecules that was designed for mimicking CA. Thus, Chapter 1 reviewed overall introduction related to CA. Chapter 2, 3 and 4 explored with respect to the especially functionalized CA mimicking catalysts, respectively. Chapter 5 summarized the previous chapters. Chapter 1 Introduction to implementation of carbonic anhydrase model complexes in carbon capture process This chapter reviewed the general CCS processes at first. In particular, this chapter focused on a biocatalyst to replace the chemical absorbents (such as amines) in the post-combustion CO2 capture. Also, CA among the biocatalyst and the previously reported CA mimicking catalysts sufficiently reviewed in order to apply to the development of following studies. Chapter 2 Kinetic study of catalytic CO2 hydration by carbonic anhydrase model complexes with various metal cation Each of CA mimicking complexes containing Tris(2-pyridylmethyl)amine (TPA) as a N4 ligand that have Zn2+, Ni2+ and Cu2+ was synthesized and characterized, which measured the effect on CO2 hydration rate according to the type of metal ion, respectively. At first, the pKa that represent intrinsic proton donating ability of water molecule bound each complex identify a tendency to increase in order of [(TPA)Cu(OH2)] < [(TPA)Ni(OH2)] < [(TPA)]Zn(OH2)] (pKa values of [(TPA)Cu(OH2)], [TPA]Ni(OH2)] and [TPA]Zn(OH2)] are 6.0, 7.6 and 8.0, respectively). Probably, this trend results from the difference in electronegativity of metal ions. The catalytic rate constants (kobs) on CO2 hydration reaction using stopped-flow spectrophotometry show a tendency to increase in order of [(TPA)Ni(OH2)] < [(TPA)Cu(OH2)] < [(TPA)Zn(OH2)]. kobs values of [(TPA)Ni(OH2)], [(TPA)Cu(OH2)] and [(TPA)Zn(OH2)] are 526.4 < 542.3 < 645.7, respectively. Although [(TPA)Ni(OH2)] is easy to deprotonate water molecule, substitution reaction (like bicarbonate release step) of bicarbonate by water molecule is not easy. But, [(TPA)Zn(OH2)] being the highest pKa value showed the fastest CO2 hydration rate. Thus, Zn2+ of CA showed that advantage of bicarbonate release step is greater than advantage of deprotonation step of water molecule. Chapter 3 Synthesis and catalytic reactivity of a Zn(II) complex, mimicking second coordination environment of CA enzyme The zinc complex using 6-(((pyridin-2-ylmethyl)(pyridin-3-ylmethyl)amino)methyl) pyridin -2-ol ligand (TPA-OH) mimicking Thr-199 existing in second coordination sphere of the CA was synthesized. The [(TPA-OH)Zn(OH2)] was investigated in comparison with [(TPA)Zn(OH2)]. The pKa value was measured by potentiometric pH titration method in order to determine the acidity of the [(TPA-OH)Zn(OH2)]. The pKa value of [(TPA-OH)Zn(OH2)] and [(TPA)Zn(OH2)] was 6.8 and 8.0, respectively. The CO2 hydration rate of [(TPA-OH)Zn(OH2)] and [(TPA)Zn(OH2)] was measured by stopped flow spectrophotometer and the measured rate constants is 648.4 and 730.6 M-1•s-1, respectively. The [(TPA-OH)Zn(OH2)] exists in intramolecular hydrogen bond network through mimicking Thr-199, so the [(TPA-OH)Zn(OH2)] showed a lower pKa value than the [(TPA)Zn(OH2)]. In addition, CO2 hydration of [(TPA-OH)Zn(OH2)] rate was also found to increase. Chapter 4 Experimental investigations on nucleophilic reaction by diverse carbonic anhydrase model Zn(II) complex with Tris(2-benzimidazolylmethyl-4-hydroxy) amine derivatives This study synthesized CA model complex using Tris(2-benzimidazolymethyl)amine derivative ligand (TBA) similar to environment around zinc ion of CA. [(TBA)Zn(OH2)] using Tris(2-benzimidazolymethyl)amine ligand mimic CA active site. [(HTBA)Zn(OH2)] using Tris(2-benzimidazolylmethyl-4-hydroxy)amine (HTBA) ligand mimic CA active site and Thr-199 existing in second coordination sphere to make hydrogen bond interaction around zinc bound water. [(STBA)Zn(OH2)] using Tris(2-benzimidazolylmethyl-6-sulfonic acid)amine (STBA) ligand and [(NTBA)Zn(OH2)] using Tris(2-benzimidazolylmethyl-6-nitro)amine (NTBA) ligand were synthesized to investigate the effect about the functional change of CA via change of electronic property of ligand. All of complexes measure the kinetic of ester hydrolysis using p-NPA to determine the ability of nucleophilic attack and catalytic efficiency. As the Km value of [(HTBA)Zn(OH2)] is 2.61 mM, the complex showed higher constant than 2.39 mM of [(TBA)Zn(OH2)] in comparison with ester hydrolysis rate. The kcat value of [(HTBA)Zn(OH2)] and [(TBA)Zn(OH2)] showed 13.48 M-1•s-1 and 10.06 M-1•s-1, respectively. The result of Km value is to prevent the formation of enzyme-substrate complex by interference of access of substrate into zinc bound hydroxide because hydroxyl group in [(HTBA)Zn(OH2)] is located at the entrance of the active site. But, kcat / Km value of [(HTBA)Zn(OH2)] have excelled rather than [(TBA)Zn(OH2)]. This result can occur easily bicarbonate release due to destabilization by electrostatic repulsion between hydroxyl group and carboxyl group produced by ester hydrolysis reaction. In comparison with ester hydrolysis kinetic of [(NTBA)Zn(OH2)], [(STBA)Zn(OH2)] and [(TBA)Zn(OH2)], [(NTBA)Zn(OH2)] showed the higher Km value than 2.37 mM of [(STBA)Zn(OH2)] as [(NTBA)Zn(OH2)] is 2.16 mM,. kcat / Km value of [(NTBA)Zn(OH2)] is 17.76 M-1•s-1 and [(STBA)Zn(OH2)] is 15.16 M-1•s-1. [(STBA)Zn(OH2)] and [(NTBA)Zn(OH2)] having electron withdrawing group such as sulfonyl group and nitro group were found that enzyme-substrate complex is more easily formed because Km value of [(TBA)Zn(OH2)] than [(STBA)Zn(OH2)] and [(NTBA)Zn(OH2)] is low, which easily formed enzyme-substrate complex when nitro group being the stronger electron withdrawing than sulfonyl group was introduced. kcat / Km value of [(STBA)Zn(OH2)] and [(NTBA)Zn(OH2)] than [(TBA)Zn(OH2)] showed the more excellent catalytic efficiency. Chapter 5 overall conclusions This chapter summarized previous chapters. And the results can provide qualitative insights for the design of improved small molecule as CO2 capture catalysts. Key word: Carbon dioxide, Carbonic anhydrase, hydration, Zinc complex, CCS, model complex, kinetic