Table 6 Selection of mutations across PET and MHET hydrolase scaffolds that resulted in an increase or decrease in enzyme activity and thermostability

From: Recent advances in enzyme engineering for improved deconstruction of poly(ethylene terephthalate) (PET) plastics

Scaffold Name

Detrimental Mutations

Beneficial Mutations

Conditions / Substrate

Citation

Activity

Thermostability

Activity

Thermostability

IsPETase

R100E, T113P, R123V, P181A, E204K, G254V

A74T, A82V, Q91P, A183T, A215P, T286V

V134L, A152S, S166T, T198V

T77E, Q119Y, W159H, I168R, D186H, S188Q, S214H, P181A

200 µL PET nanoparticles, 10 µL of 0.1 mg/mL enzyme stock, 37 °C, 1 h, 300 µL rxn, pH 9

Cui, et al.90

I168R/D186H/S214H

I168R/S214H, D186H/S214H, W159H/I168R/S188Q/S214H

LCC

T96M, Y127G, D238C/S283C, D238C/F243W/N246D/S283C

F243I

F243I/W, N246D

T96M, Y127G, N246D/M, D238C/S283C

100 mg preform PET, 1 mL of 0.69 µM enzyme stock, 70 °C, 100 mL rxn, pH 8

Tournier, et al.29

D238C/F243I/N246D/S283C

D238C/F243I/S283C, Y127G/D238C/F243I/S283C, D238C/F243I/N246M/S283C

PHL7

F63Y/A, M132W, L210F

L210V/I, L93A

L210T/V, L93A

F63Y/A, Q95G, M132W, L210F

~45 mg amorphous PET coupons, 0.55 mg enzyme/g PET, 70 °C, 1.8 mL rxn, 16 h, pH 8

Richter, et al.50

H130W, H185S, F189I

L93F, Q95Y, L210T/A/S, D233K

IsMHETase

S225A, R411A/Q, 416 A/G, F424Q/N, D492A, H528A, R411A/S416A, R411A/S416G, R411A/S419G, R411Q/S416A, R411Q/S416G

n.d.

W297A, F415A

n.d.

30 min, 30 °C, 6 nM enzyme, 1 mM MHET, pH 7.5

Palm, et al.144

  1. The list is not exhaustive. One reference which disclosed thermostability and activity data for single and multiple mutation variants was chosen for each scaffold. Mutations (single or multiple) comprising a variant were classified whether or not they improved or decreased the activity upon comparison to the wild-type scaffold under the tested conditions. For the PET hydrolase scaffolds, IsPETase, LCC, and PHL7, activity refers to PET hydrolysis, while for IsMHETase, activity refers to MHET hydrolysis. Thermostability of variants was not measured by Palm, et al.144, so the corresponding column is marked “n.d.” for not determined.
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