Altering lipase activity and enantioselectivity in organic media using organo‐soluble bases: Implication for rate‐limiting proton transfer in acylation step

Abstract

With the hydrolytic resolution of (R,S)-naproxen 2,2,2-trifluoroethyl esters via a partially purified papaya lipase (PCPL) in water-saturated isooctane as the model system, the enzyme activity, and enantioselectivty is altered by adding a variety of organo-soluble bases that act as either enzyme activators (i.e., TEA, MP, TOA, DPA, PY, and DMA) or enzyme inhibitors (i.e., PDP, DMAP, and PP). Triethylamine (TEA) is selected as the best enzyme activator as 2.24-fold increase of the initial rate for the (S)-ester is obtained when adding 120 mM of the base. By using an expanded Michaelis-Menten mechanism for the acylation step, the kinetic analysis indicates that the proton transfer for the breakdown of tetrahedral intermediates to acyl-enzyme intermediates is the rate-limiting step, or more sensitive than that for the formation of tetrahedral intermediates when the enzyme activators of different pKa are added. However, no correlation for the proton transfers in the acylation step is found when adding the bases acting as enzyme deactivators.

Kinetic modeling of lipase‐catalyzed esterification reaction between oleic acid and trimethylolpropane: A simplified model for multi‐substrate multi‐product ping–pong mechanisms

“…Considering that OA is a fatty acid with long carbon chain that is a rather bulky acyl donor substrate for CALB, the enzyme acylation including the breakdown of the tetrahedral intermediate and the side‐product release together with the formation of a covalent acyl‐enzyme complex is assumed to be much slower than the deacylation by a nucleophile ( k 4 , k 6 and k 8 >> k 2 ) and is controlled by the proton transfer rate within the catalytic triad for formation of the acyl‐enzyme . Therefore the limiting rates shown in Table can be approximated equal ( V 1 * ≈ V 2 * ≈ V 3 * ).…”

Mixed systems to assist enzymatic ring opening polymerization of lactide stereoisomers

“…Apart from enzymatic polymerization, some authors studied the inuence of various organic bases (aromatic tertiary amine with pyridine and some pyridine derivatives, and aliphatic tertiary amine like triethylamine, N-methylpiperidine) on lipases activity towards the hydrolytic resolution of esters reporting signicant lipase activation with some of the tested bases. 32 Interestingly, to the best of our knowledge, such activation of lipase by organic bases has not been tested and reported so far for lactide eROP.…”

Lipase‐catalyzed dynamic hydrolytic resolution of (R,S)‐2,2,2‐trifluoroethyl α‐chlorophenyl acetate in water‐saturated isooctane

“…To our surprise, a 38.3‐fold enhancement of A R at ( B ) = 80 mmol dm −3 is obtainable. This is more than an order of magnitude higher than values found for the hydrolysis of ( R , S )‐2,2,2‐trifluoroethyl naproxen ester at 45 °C 21. More studies on using an expanded Michälis–Menten kinetics for the acylation step are needed in order to elucidate whether the base effect is increasing the substrate affinity for the active site or proton transfer for the formation or breaking of the tetrahedral intermediate of the acylation step 21…”