Oxidoreductases articles from across Nature Portfolio

It can be challenging to obtain meaningful and accurate structural information for air-sensitive proteins. This protocol describes the application of customized vacuum manifold and anaerobic chamber setups for the purification and cryo-electron microscopy analysis of air-sensitive nitrogenase enzymes.

Here, the authors characterise an aldo-keto reductase AKRtyl, which belongs to a previously unidentified subfamily AKR12D. They uncover a complex mechanism of allosteric regulation that is mediated by 3 distinct states.

Understanding the complex relationships between enzyme sequence, folding stability and catalytic activity is essential for applications, but current technologies cannot simultaneously resolve both stability and activity phenotypes and couple these to gene sequences at large scale. Here, the authors report Enzyme Proximity Sequencing (EP-Seq), a deep mutational scanning method to assay both expression level and catalytic activity of thousands of oxidoreductase variants from a cellular pool in a single experiment.

Ipatasertib is a potent Akt (protein kinase B) inhibitor synthesized via a chemoenzymatic process. Here, the authors use mutational scanning and algorithm-aided enzyme engineering to optimize a ketoreductase from Sporidiobolus salmonicolor and generate a 10-amino acid substituted variant exhibiting a 64-fold higher k_cat and improved yield for the relevant alcohol intermediate, with ≥ 98% conversion and a diastereomeric excess of 99.7% (R,R-trans) from 100 g L⁻¹ ketone after 30 h.

Fusion proteins between roGFP2 and glutaredoxins are used for intracellular redox measurements. Here, the authors determined all rate constants of the reaction cycle for roGFP2 measurements and identified an alternative glutaredoxin mechanism.

Converting simple building blocks into enantiopure amines is an important synthetic transformation. Now, a biocatalyst has been discovered that catalyses two reductive steps within a single active site, and can synthesize a range of amine targets from ene–one starting materials.

Nitrogenase reduces dinitrogen at one of its iron–sulfur cores to produce ammonia by a convoluted mechanism. Now, research highlights the importance of sulfur mobility on one of nitrogenase’s metallocofactors for nitrogen fixation.

Levine and colleagues demonstrate that the effects of caloric restriction on hepatic gene expression, circulating acylcarnitines and body temperature are largely mediated by an increase in the daytime peak of NADH concentration in the liver.

Conventional experiments for generating proteins with improved properties by directed evolution are iterative, lengthy and costly. Now, a label-free assay has been developed for ultrahigh-throughput microfluidic screening that can dramatically accelerate the discovery of superior biocatalysts from a single round of genetic randomization.