Met80 of cytochrome c (cyt c) has been shown to dissociate from its heme iron when cyt c interacts with cardiolipin (CL), which triggers the release of cyt c into the cytosol initiating apoptosis. We found that the mass of human cyt c increases by 16 Da in the Met80-Lys86 region by reaction with molecular oxygen in the presence of CL-containing liposomes and dithiothreitol (DTT). To investigate the effect of Met80 dissociation on the reaction of cyt c with molecular oxygen without affecting its secondary structures, a human cyt c mutant (Δ8384 cyt c) was constructed by removing two amino acids (Val83 and Gly84) from the loop containing Met80. According to MALDI-TOF-MS and tandem mass measurements, Met80 of Δ8384 cyt c was modified site-specifically to methionine sulfoxide when purified in the presence of molecular oxygen, whereas Met80 was not modified in the absence of molecular oxygen. A red-shift of the Soret band from 406 to 412 nm and absorption increase at ∼536 and ∼568 nm were observed for Δ8384 cyt c when it reacted with DTT and molecular oxygen, followed by a further red-shift of the Soret band to 416 nm and absorption increase at ∼620 and ∼650 nm. These results indicate that Met80 of cyt c is oxidized site-specifically by formation of the oxy and subsequent compound I-like species when Met80 dissociates from the heme iron, where the Met80 modification may affect its peroxidase activity related to apoptosis.

The present study aims to design 3D scaffold hydroxyapatite (HA)/collagen (Coll) based egg‐white (EW) as antibacterial properties. The calcium source in HA synthesis derived from the Pinctada maxima shell cultivated on Bali Island has proven biocompatibility, and the compressive strength exceeded human bone. HA synthesis by precipitation with heat treatment in oven‐dried at 80°C (HA‐80) and annealed at 900°C (HA‐900), has crystallinity 48% and 85%, respectively, were used for scaffold design. The physicochemical properties of X‐ray diffractometer spectra showed that increasing temperature affected the crystallinity and HA phase formed. Furthermore, the crystal structure of HA changed in nanocomposite due to the substitution of Coll and EW, and the Fourier transform infrared spectroscopy spectra confirmed that the absorption peak of the phosphate group (1027–1029 cm−1) decreased intensity, presumably by protein binding of EW and Coll. The cell viability of HA/Coll/EW in 24, 48, and 72 h incubation period was 112.34 ± 4.36, 104.89 ± 3.41, 72.88 ± 6.85, respectively. The decreases of cell viability due to high cell density and reduced nutrients in wells. Antibacterial activity of HA/Col/EW exhibited a strong zone of inhibition against bacteria causing periodontitis; Porphyromonas gingivalis, Aggregatibacter actinomycetemcomitans, Fusobacterium nucleatum, and Staphylococcus aureus.

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We have previously shown that methionine-heme iron coordination is perturbed in domain-swapped dimeric horse cytochrome c. To gain insight into the effect of methionine dissociation in dimeric cytochrome c, we investigated its interaction with cyanide ion. We found that the Soret and Q bands of oxidized dimeric cytochrome c at 406.5 and 529 nm redshift to 413 and 536 nm, respectively, on addition of 1 mM cyanide ion. The binding constant of dimeric cytochrome c and cyanide ion was obtained as 2.5 × 10(4) M(-1). The Fe-CN and C-N stretching (ν (Fe-CN) and ν (CN)) resonance Raman bands of CN(-)-bound dimeric cytochrome c were observed at 443 and 2,126 cm(-1), respectively. The ν (Fe-CN) frequency of dimeric cytochrome c was relatively low compared with that of other CN(-)-bound heme proteins, and a relatively strong coupling between the Fe-C-N bending and porphyrin vibrations was observed in the 350-450-cm(-1) region. The low ν (Fe-CN) frequency suggests weaker binding of the cyanide ion to dimeric cytochrome c compared with other heme proteins possessing a distal heme cavity. Although the secondary structure of dimeric cytochrome c did not change on addition of cyanide ion according to circular dichroism measurements, the dimer dissociation rate at 45 °C increased from (8.9 ± 0.7) × 10(-6) to (3.8 ± 0.2) × 10(-5) s(-1), with a decrease of about 2 °C in its dissociation temperature obtained with differential scanning calorimetry. The results show that diatomic ligands may bind to the heme iron of dimeric cytochrome c and affect its stability.

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