The treatment of environmental pollutants such as synthetic dyes and lignin has received much attention, especially for biotechnological treatments using both native and artificial metalloenzymes. In this study, we designed and engineered an efficient peroxidase using the O₂ carrier myoglobin (Mb) as a protein scaffold by four mutations (F43Y/T67R/P88W/F138W), which combines the key structural features of natural peroxidases such as the presence of a conserved His-Arg pair and Tyr/Trp residues close to the heme active center. Kinetic studies revealed that the quadruple mutant exhibits considerably enhanced peroxidase activity, with the catalytic efficiency (k_cat/K_m) comparable to that of the most efficient natural enzyme, horseradish peroxidase (HRP). Moreover, the designed enzyme can effectively decolorize a variety of synthetic organic dyes and catalyze the bioconversion of lignin, such as Kraft lignin and a model compound, guaiacylglycerol-β-guaiacyl ether (GGE). As analyzed by HPLC and ESI-MS, we identified several bioconversion products of GGE, as produced via bond cleavage followed by dimerization or trimerization, which illustrates the mechanism for lignin bioconversion. This study indicates that the designed enzyme could be exploited for the decolorization of textile wastewater contaminated with various dyes, as well as for the bioconversion of lignin to produce more value-added products.

Protein glycation is an important protein post-translational modification and is one of the main pathogenesis of diabetic angiopathy. Other than glycated hemoglobin, the protein glycation of other globins such as myoglobin (Mb) is less studied. The protein glycation of human Mb with ribose has not been reported, and the glycation sites in the Mb remain unknown. This article reports that d-ribose undergoes rapid protein glycation of human myoglobin (HMb) at lysine residues (K34, K87, K56, and K147) on the protein surface, as identified by ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS) and electrospray ionization tandem mass spectrometry (ESI-MS/MS). Moreover, glycation by d-ribose at these sites slightly decreased the rate of the met heme (Fe^III) in reaction with H₂O₂ to form a ferryl heme (Fe^IV=O). This study provides valuable insight into the protein glycation by d-ribose and provides a foundation for studying the structure and function of glycated heme proteins.

Imbalance in the cellular redox system is thought to be associated with the induction and progression of breast cancers, and heme proteins may regulate the redox balance. Cytochrome b₅ (Cyt b₅) is a small mitochondrial heme protein. Its function and regulating mechanism in breast cancer remain unknown. In this study, we elucidated the level of endogenous oxidative stress in breast cancer cells, MCF-7 cells (hormone receptor-positive cells) and MDA-MB-231 cells (triple-negative cells), and investigated the difference in Cyt b₅ content. Based on the low content of Cyt b₅ in MDA-MB-231 cells, the overexpression of Cyt b₅ was found to regulate the oxidative stress and apoptosis cascades, including ERK1/2 and Akt signaling pathways. The overexpressed Cyt b₅ MDA-MB-231 cells were shown to exhibit decreased oxidative stress, less phosphorylation of ERK1/2 and Akt, and less cleavage of caspases 3 and 9 upon treatment with H₂O₂, as compared to those of normal MDA-MB-231 cells. Moreover, the overexpressed Cyt b₅ most likely functioned by interacting with its protein partner, Cyt c, as suggested by co-immunoprecipitation studies. These results indicated that Cyt b₅ has different effects on breast cancer cells of different phenotypes, which provides useful information for understanding the multiple roles of Cyt b₅ and provides clues for clinical treatment.