Characterization of PPS Piston and Packing Ring Materials for High-Pressure Hydrogen Applications.

The widespread adoption of renewable energy hinges on the efficient transportation of hydrogen. Reciprocating piston compressor technology in non-lubricated operation will play a key role, ensuring high flow rates and compression ratios. These systems rely on advanced high-strength sealing solutions for piston and rod packing rings utilizing advanced fiber-reinforced polymers.

Influence of cysteine 164 on active site structure in rat cysteine dioxygenase.

Cysteine dioxygenase is a non-heme mononuclear iron enzyme with unique structural features, namely an intramolecular thioether cross-link between cysteine 93 and tyrosine 157, and a disulfide bond between substrate L-cysteine and cysteine 164 in the entrance channel to the active site. We investigated how these posttranslational modifications affect catalysis through a kinetic, crystallographic and computational study. The enzyme kinetics of a C164S variant are identical to WT, indicating that disulfide formation at C164 does not significantly impair access to the active site at physiological pH.

The cysteine dioxygenase homologue from Pseudomonas aeruginosa is a 3-mercaptopropionate dioxygenase.

Thiol dioxygenation is the initial oxidation step that commits a thiol to important catabolic or biosynthetic pathways. The reaction is catalyzed by a family of specific non-heme mononuclear iron proteins each of which is reported to react efficiently with only one substrate. This family of enzymes includes cysteine dioxygenase, cysteamine dioxygenase, mercaptosuccinate dioxygenase, and 3-mercaptopropionate dioxygenase.

Correlating crosslink formation with enzymatic activity in cysteine dioxygenase.

Cysteine dioxygenase (CDO) from rat and other mammals exhibits a covalent post-translational modification between the residues C93 and Y157 that is in close proximity to the active site, and whose presence enhances the enzyme's activity. Protein with and without C93-Y157 crosslink migrates as distinct bands in SDS-PAGE, allowing quantification of the relative ratios between the two forms by densitometry of the respective bands. Expression of recombinant rat wild type CDO in Escherichia coli typically produces 40-50% with the C93-Y157 crosslink.

Simplified cysteine dioxygenase activity assay allows simultaneous quantitation of both substrate and product.

A high-performance liquid chromatography (HPLC) method for enzyme activity assays using a hydrophilic interaction liquid chromatography (HILIC) column in combination with an evaporative light scattering detector was developed. The method was used to measure the activity of the non-heme mono-iron enzyme cysteine dioxygenase. The substrate cysteine and the product cysteine sulfinic acid are very weak chromophores, making direct ultraviolet (UV) detection without derivatization rather insensitive; moreover, derivatization of cysteine is often not efficient.