Dequan Zhang

Effects of protein phosphorylation on color stability of ground meat.

The influence of protein phosphorylation on meat color stability was investigated in this study. Phosphatase and protein kinase inhibitors were added to minced ovine Longissimus thoracis et lumborum (LTL) muscle to manipulate the global phosphorylation of sarcoplasmic proteins. The data obtained show that the rate and extent of pH decline, along with lactate accumulation in postmortem muscle, were related to protein phosphorylation. Analysis of meat color and the relative content of myoglobin redox forms revealed that meat color stability was inversely related to the phosphorylation of sarcoplasmic proteins. Thus, this study suggests that protein phosphorylation may be involved in meat color development by regulating glycolysis and the redox stability of myoglobin.

Phosphorylation prevents in vitro myofibrillar proteins degradation by μ-calpain.

Myofibrillar proteins degradation contributes to meat tenderisation during post-mortem ageing. Protein phosphorylation has been revealed to be associated with meat tenderness in recent years. This study was undertaken to determine the impact of myofibrillar proteins phosphorylation on the degradation susceptibility by μ-calpain. Myofibrillar proteins were first incubated with protein kinase A (PKA) or alkaline phosphatase (AP) to increase or decrease the phosphorylation level, following μ-calpain hydrolysis. Myosin heavy chain, actin, desmin and troponin T showed different levels of degradation in control, AP and PKA groups under different Ca2+ concentrations. Generally, more degradation products were detected with the increase of Ca2+ concentration. Compared to the control, the protein degradation was higher in AP-treated group and lower in PKA-treated group. This study shows that phosphorylation prevents proteolytic susceptibility of myofibrillar proteins to degradation by μ-calpain, indicating that protein phosphorylation plays an important role in meat tenderisation during post-mortem ageing.

LF-NMR to explore water migration and water–protein interaction of lamb meat being air-dried at 35°C

ABSTRACT To decrease the drying time and energy consuming, water migration and water–protein interactions of lamb meat being air-dried at 35°C were investigated. Low-field nuclear magnetic resonance T2b indicated the water–protein interactions changed during air-drying. Area of T21 (intramyofibrillar water) decreased, meanwhile the area of T22 (extra-myofibrillar water) increased when the moisture content decreased from 55 to 45%, indicating the water migrated from myofibril to extramyofibril. Drying rate (the rate of water migration from meat to air) could be predicted by the area of T2 populations, and the correlation coefficient was 0.990. Change of water binding and hydration in myofibrils was evident by the increase in hydrophobicity and decrease in solubility of myofibrillar protein. Differential scanning calorimetry showed denaturation of myosin in dried meat which might result in water migration from myofibril to extramyofibril space. In conclusion, water–protein interactions changed, and then influenced the drying rate during drying.

Comparative profiling of sarcoplasmic phosphoproteins in ovine muscle with different color stability

The phosphorylation of sarcoplasmic proteins in postmortem muscles was investigated in relationship to color stability in the present study. Although no difference was observed in the global phosphorylation level of sarcoplasmic proteins, difference was determined in the phosphorylation levels of individual protein bands from muscles with different color stability. Correlation analysis and liquid chromatography - tandem mass spectrometry (LC-MS/MS) identification of phosphoproteins showed that most of the color stability-related proteins were glycolytic enzymes. Interestingly, the phosphorylation level of myoglobin was inversely related to meat color stability. As the phosphorylation of myoglobin increased, color stability based on a∗ value decreased and metMb content increased. In summary, the study revealed that protein phosphorylation might play a role in the regulation of meat color stability probably by regulating glycolysis and the redox stability of myoglobin, which might be affected by the phosphorylation of myoglobin.

Antemortem stress regulates protein acetylation and glycolysis in postmortem muscle.

Although exhaustive research has established that preslaughter stress is a major factor contributing to pale, soft, exudative (PSE) meat, questions remain regarding the biochemistry of postmortem glycolysis. In this study, the influence of preslaughter stress on protein acetylation in relationship to glycolysis was studied. The data show that antemortem swimming significantly enhanced glycolysis and the total acetylated proteins in postmortem longissimus dorsi (LD) muscle of mice. Inhibition of protein acetylation by histone acetyltransferase (HAT) inhibitors eliminated stress induced increase in glycolysis. Inversely, antemortem injection of histone deacetylase (HDAC) inhibitors, trichostatin A (TSA) and nicotinamide (NAM), further increased protein acetylation early postmortem and the glycolysis. These data provide new insight into the biochemistry of postmortem glycolysis by showing that protein acetylation regulates glycolysis, which may participate in the regulation of preslaughter stress on glycolysis in postmortem muscle.

Phosphorylation regulated by protein kinase A and alkaline phosphatase play positive roles in μ-calpain activity

This study was aimed to determine the effect of phosphorylation/dephosphorylation regulated by protein kinase A (PKA) and alkaline phosphatase (AP) on μ-calpain activity at different Ca2+ concentrations. μ-Calpain was treated with AP or PKA at 0.01, 0.05, 0.1 and 1 mM Ca2+. The pH value decreased in the AP group but remained stable in the control and PKA groups during incubation. Except samples incubated at 0.01 and 0.1 mM Ca2+ for more than 20 min, μ-calpain incubated with PKA showed a higher degree of autolysis than control, but lower than the AP group. The content of α-helix structure of μ-calpain increased as phosphorylation level rose. Phosphorylation of μ-calpain at serine 255, 256, 476, 417 and 420 was identified. PKA catalyzed μ-calpain phosphorylation at serine 255, 256 and 476, located at domains II and III, positively regulated μ-calpain activity. These data demonstrated that dephosphorylation and PKA phosphorylation positively regulated μ-calpain activity, which was limited by increased Ca2+ concentration.

Dephosphorylation enhances postmortem degradation of myofibrillar proteins

Protein degradation is primarily responsible for postmortem meat tenderization, which might be affected by phosphorylation. The objective of this study was to investigate the effect of phosphorylation on myofibrillar proteins degradation in muscle during postmortem. Here we modulated the phosphorylation status of protein by protein kinase inhibitor and phosphatase inhibitor, and the effect of these inhibitors on myofibrillar protein degradation was evaluated. Generally, myofibril fragmentation index of samples with lower phosphorylation level was higher. Troponin T and heat shock protein 27 were degraded faster in protein kinase inhibited (low phosphorylation level) muscle, compared with the other two groups, while the degradation of desmin was not affected by inhibitors. Meanwhile, myosin heavy chain, actin and tropomyosin showed limited degradation in postmortem muscle. This study showed that dephosphorylation enhances the degradation of some myofibrillar proteins, indicating that protein phosphorylation may play an important role in postmortem meat tenderization.

Calpastatin inhibits the activity of phosphorylated μ-calpain in vitro

The objective of this study was to investigate the effect of phosphorylation on the sensitivity of μ-calpain to the inhibition induced by calpastatin. Purified μ-calpain was incubated with alkaline phosphatase (AP) or protein kinase A (PKA) to modulate the phosphorylation level of μ-calpain. Accurately 25, 50, 100 and 150 units of AP/PKA-treated μ-calpain were mixed with the same amounts of heat stable proteins and incubated at 4 °C. In the calpastatin-free system, AP and PKA-treated μ-calpain had higher proteolytic activity compared to the control. Intact AP-treated μ-calpain degraded fastest in the 50, 100 and 150 unit μ-calpain incubation systems. However, the degradation rate of μ-calpain in control and PKA group was non-significant in 100 and 150 unit μ-calpain systems. Our results demonstrated that, compared to dephosphorylated and control μ-calpain, calpastatin presents greater inhibition to PKA phosphorylated μ-calpain. This study increases understanding of the mechanism of μ-calpain activity regulated by phosphorylation.

Postmortem ageing influences the thawed meat quality of frozen lamb loins

The aim of this study was to determine the effect of postmortem ageing on quality attributes of frozen/thawed lamb loins. The loins (M. Longissimus dorsi; n = 30) were randomly divided into six groups: five frozen treatment groups and the control (4 °C for 120 h). Treatment groups were frozen for 3 weeks and thawed at 4 °C overnight, muscles were preserved at 4 °C until 120 h. Compared to the control, pH values and color of frozen meat declined (P < 0.05) after ageing for 72 h, lower shear force values and higher water loss with earlier freezing were due to extensive degradation of myofibril protein. These results indicated the loins frozen in earlier period could accelerate ageing rate, but impaired meat quality inevitably, freezing between 12 h and 24 h postmortem was a better consideration.

Meat Color Stability of Ovine Muscles Is Related to Glycolytic Dehydrogenase Activities: Meat color stability of ovine muscles…

The relationship between glycolytic dehydrogenase, including glyceraldehyde-3-phosphate dehydrogenase (GAPDH), lactic dehydrogenase (LDH), and meat color stability was studied in this study using ovine muscle. Three different ovine muscles, including M. longissimus lumborum (LL), M. semimembranosus (SM), and M. psoas major (PM), were obtained (n = 10, respectively), and then displayed for 7 days at 4 °C. The LL and SM muscle had higher surface redness, higher (P < 0.05) GAPDH activity, nicotinamide adenine dinucleotide (NADH) content, and lower (P < 0.05) LDH-B activity than PM muscles during display. The PM muscle had the worst color stability and lowest NADH content. These results suggest that variation in color stability of physiologically different muscles may be affected by glycolysis dehydrogenases. Comparatively, our data showed that GAPDH may play a more important role than LDH-B to maintain meat color stability.