Interactions between unfolding/disassembling behaviors, proteolytic subfragments and reversible aggregation of oxidized skeletal myosin isoforms at different salt contents.

Myosin filament plays a critical role in water-trapping and thermodynamic regulation during processing of brined muscle foods. The redox state and availability of proteolytic/antioxidant enzymes affected by salt may change the ion-binding capacity of myosin consequently contributing to swelling and rehydration. Thus, this study investigated the impact of different salt content (0%, 1%, 2%, 3%, 4%, 5% NaCl) and oxidation in vitro (10 mM H2O2/ascorbate-based hydroxyl radical (OH)-generating system) on the oxidative stability, solubility/dispersion capacity, chymotrypsin digestibility, aggregation site and the microrheological properties of isolated porcine myosin. The result showed that, brining at 2% salt exposed more sulfhydryl groups and inhibited the formation of disulfide bond, whereby smaller dispersed structure (diameter within 10-50 nm) and higher Ca2+-ATPase activity of the denatured myosin were observed. Accordingly, gel electrophoresis showed that myosin S1 and HMM subunits were highly oxidized and susceptible to reversible assembles. Despite enhanced hydrophobic interactions between swelled myosin at 3% salt content, ≥4% salt greatly promoted the exposure/polarization of tryptophan and cross-linking structures, mainly occurring at myosin S2 portion. The results of micro-rheology proved that oxidized myosin formed a tighter heat-set network following rehydration at high ion strength (≥4% salt), suggesting an increased inter-droplet resistance and macroscopic viscosity. This work is expected to give some useful insights into improved texture and functionality of engineered muscle foods.

Promotion effect of salt on intramuscular neutral lipid hydrolysis during dry-salting process of porcine (biceps femoris) muscles by inducing phosphorylation of ATGL, HSL and their regulatory proteins of Perilipin1, ABHD5 and G0S2.

Towards a better understanding of the formation mechanism of salt on intramuscular triglyceride (TG) hydrolysis occurring in biceps femoris (BF) muscles during dry-salting process, the changes of TG hydrolysis, TG hydrolysis activity and phosphorylation of adipose triglyceride lipase (ATGL) and hormone sensitive lipase (HSL) as well as their regulatory proteins (Perilipin1, ABHD5, G0S2) with different salt content (0%, 1%, 3%, 5%) and salting time (the first and third day) were analyzed. The results showed that dry-salting significantly increased the TG hydrolase activity and hydrolysis extent with salting process proceed (P < 0.05), especially upon the treatment with 3% amount of salt. The SDS-PAGE and Western-blot results further demonstrated that the promotion of salt on TG hydrolysis in intramuscular adipocytes was mainly attributed to the activation of protein kinase activity and protein phosphorylation process. Accordingly, the ATGL and HSL were activated, and meanwhile, the TG hydrolysis pivotal switch perilipin1 was also turned on by phosphorylation modification.

Impact of salt content and hydrogen peroxide-induced oxidative stress on protein oxidation, conformational/morphological changes, and micro-rheological properties of porcine myofibrillar proteins.

Salting and rehydration of myofibrils can be interfered with free radical diffusion process. This study investigated the effects of salt content (0, 1, 3 and 5%) and H2O2/ascorbate-based hydroxyl radical (OH)-generating system (1, 10, 20 mM H2O2) on the oxidation, conformation, aggregation, and thermal stability of porcine myofibrillar proteins (MPs). Results showed that 5% of salt inhibited carbonylation of MPs with intensive sulfhydryl loss and tryptophan quenching. Fourier transform infrared (FTIR), laser light scattering, and scanning electron microscopy (SEM) suggested that 20 mM H2O2 transformed more α-helix into ß-sheet of MPs, favoring larger aggregates being selectively exposed towards solvent during salt-induced fiber swelling. Oxidized MPs brined with ≤1% salt underwent partial unfolding with higher flexibility, while up to 5% of salt greatly hampered their hydration potential and weakened inter-fibrillar hydrogen bond with an improved protein solubility. Micro-rheology revealed that 1% of salt and 10 mM H2O2 rendered a denser structure of heat-set MPs gels.