Myoglobin Post-Translational Modifications Influence Color Stability of Beef Longissimus Lumborum

Post-translational modifications (PTM) of proteins play critical roles in biological processes. PTM of muscle proteins influence meat quality. Nonetheless, myoglobin (Mb) PTM and their impact on fresh beef color stability have not been characterized yet. Therefore, our objectives were to identify Mb PTM in beef longissimus lumborum muscle during postmortem aging and to characterize their influence on color stability. The longissimus lumborum muscles from 9 (n = 9) beef carcasses (24 h postmortem) were subjected to wet aging for 0, 7, 14, and 21 d. At the end of each wet-aging period, steaks were fabricated. One steak for analyses of PTM was immediately frozen at −80°C, whereas other steaks were assigned to refrigerated storage in the darkness under aerobic packaging. Instrumental color and biochemical attributes were evaluated on day 0, 3, or 6 of storage. Mb PTM were analyzed using two-dimensional electrophoresis and tandem mass spectrometry. Surface redness (a* value), color stability, and Mb concentration decreased (P < 0.05) upon aging. Gel image analyses identified 6 Mb spots with similar molecular weight (17 kDa) but different isoelectric pH. Tandem mass spectrometry identified multiple PTM (phosphorylation, methylation, carboxymethylation, acetylation, and 4-hydroxynonenal alkylation) in these 6 isoforms. The amino acids susceptible to phosphorylation were serine (S), threonine (T), and tyrosine, whereas other PTM were detected in lysine (K), arginine (R), and histidine residues. Additionally, distal histidine (position 64), critical to heme stability, was found to be alkylated. Overall, Mb PTM increased with aging. The aginginduced PTM, especially those occurring close to hydrophobic heme pocket, could disrupt Mb tertiary structure, influence heme affinity, and compromise oxygen binding capacity, leading to decreased color stability of fresh beef. Furthermore, PTM at K45, K47, and K87 were unique to Mb from non-aged beef, whereas PTM at R31, T51, K96, K98, S121, R139, and K147 were unique to Mb from aged counterparts, indicating that these Mb PTM could be used as novel biomarkers for fresh beef color stability.