Electron paramagnetic resonance studies and magnetic properties of low symmetry niobium(IV) complexes

Antholine, W E

doi:10.31274/rtd-180815-2547

Cited by 1 publication

(1 citation statement)

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“…EPR spectral fits were performed using the SIMPOW program. 30 Low-temperature MCD spectra were recorded with a Jasco J-715 spectropolarimeter in conjunction with an Oxford Instruments SpectromagPT 7 Tesla magnetocryostat. MCD spectra are presented as the difference between spectra obtained with the magnetic field aligned parallel and antiparallel to the light propagation axis to eliminate contributions from the circular dichroism background and glass strain.…”

Section: ■ Materials and Methodsmentioning

confidence: 99%

A Single DNA Point Mutation Leads to the Formation of a Cysteine–Tyrosine Crosslink in the Cysteine Dioxygenase from Bacillus subtilis

et al. 2023

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Cysteine dioxygenase (CDO) is a non-heme ironcontaining enzyme that catalyzes the oxidation of cysteine (Cys) to cysteine sulfinic acid (CSA). Crystal structures of eukaryotic CDOs revealed the presence of an unusual crosslink between the sulfur of a cysteine residue (C93 in Mus musculus CDO, MmCDO) and a carbon atom adjacent to the phenyl group of a tyrosine residue (Y157). Formation of this crosslink occurs over time as a byproduct of catalysis and increases the catalytic efficiency of CDO by at least 10-fold. Interestingly, in bacterial CDOs, the residue corresponding to C93 is replaced by a highly conserved glycine (G82 in Bacillus subtilis CDO, BsCDO), which precludes the formation of a C−Y crosslink in these enzymes; yet bacterial CDOs achieve turnover rates paralleling those of fully crosslinked eukaryotic CDOs. In the present study, we prepared the G82C variant of BsCDO to determine if a single DNA point mutation could lead to C−Y crosslink formation in this enzyme. We used gel electrophoresis, peptide mass spectrometry, electron paramagnetic resonance spectroscopy, and kinetic assays to characterize this variant alongside the natively crosslinked wild-type (WT) MmCDO and the natively noncrosslinked WT BsCDO. Collectively, our results provide compelling evidence that the G82C BsCDO variant is indeed capable of C−Y crosslink formation. Our kinetic studies indicate that G82C BsCDO has a reduced catalytic efficiency compared to WT BsCDO and that activity increases as the ratio of crosslinked to non-crosslinked enzyme increases. Finally, by carrying out a bioinformatic analysis of the CDO family, we were able to identify a large number of putatively crosslinked bacterial CDOs, the majority of which are from Gram-negative pathogenic bacteria.

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Section: ■ Materials and Methodsmentioning

confidence: 99%