Trading off stability against activity in extremophilic aldolases

Abstract

Understanding enzyme stability and activity in extremophilic organisms is of great biotechnological interest, but many questions are still unsolved. Using 2-deoxy-D-ribose-5-phosphate aldolase (DERA) as model enzyme, we have evaluated structural and functional characteristics of different orthologs from psychrophilic, mesophilic and hyperthermophilic organisms. We present the first crystal structures of psychrophilic DERAs, revealing a dimeric organization resembling their mesophilic but not their thermophilic counterparts. Conversion into monomeric proteins showed that the native dimer interface contributes to stability only in the hyperthermophilic enzymes. Nevertheless, introduction of a disulfide bridge in the interface of a psychrophilic DERA did confer increased thermostability, suggesting a strategy for rational design of more durable enzyme variants. Constraint network analysis revealed particularly sparse interactions between the substrate pocket and its surrounding α-helices in psychrophilic DERAs, which indicates that a more flexible active center underlies their high turnover numbers.

Dimerization of Proline Dehydrogenase from Thermus thermophilus Is Crucial for Its Thermostability

“…One of the strategies to increase thermostability is to provide rigidity through oligomerization . Trade‐offs between thermal stability and activity made via a higher oligomeric species have been reported and our results confirm that oligomerization might be a common strategy to confer thermostability to an enzyme at the expense of its catalytic activity.…”

The high-energy transition state of a membrane transporter

“…These robust, 10-20 fold, increases transport rate in a set of GltPh variants that contain at most four mutations, are in line with similar efforts in enzymology [68][69][70][71][72][73][74] .…”

DERA in Flow: Synthesis of a Statin Side Chain Precursor in Continuous Flow Employing Deoxyribose-5-Phosphate Aldolase Immobilized in Alginate-Luffa Matrix

“…The major issue with aromatic substrates is the limited solubility in aqu. buffer systems, even in mixtures with DMSO [38]. These substrates would be especially interesting, as the direct addition of acetaldehyde to the molecule core would reduce the number of process steps drastically.…”

“…Acrolein was also tested as substrate. Although the mutant was successfully tested for its tolerance toward crotonaldehyde, acrolein still inhibits the enzyme [38,39]. Further investigations were conducted using acetaldehyde 1 and chloroacetaldehyde 2.…”