磷酸化水平对肌红蛋白稳定性的影响

doi:10.3864/j.issn.0578-1752.2017.22.014

Abstract

Abstract: 【Objective】Myoglobin is the main pigment responsible for meat color, which is mainly in sarcoplasm. Meat color is determined by the absolute content and the dynamics of myoglobin redox form interconversions. Studies have shown that protein phosphorylation may play a negative role in meat color development by regulating glycolysis and the redox stability of myoglobin. The effects of phosphorylation on myoglobin stability was investigated in this study, which will provide theoretical basis for improving meat color stability through regulating protein phosphorylation level.【Method】The pure metmyoglobin from skeletal muscle was used. Metmyoglobin was reduced by sodium dithionite, which was then removed by ultrafiltration. After that, alkaline phosphatase (AP) was added to catalyze the dephosphorylation of myoglobin in vitro. The phosphorylation level of myoglobin was determined by using SDS-PAGE and then stained by Pro-Q & Ruby reagent. pH value was measured by a pH meter. The relative content of three myoglobin forms (oxygenated myoglobin, deoxy myoglobin and iron myoglobin) and the secondary structure of myoglobin during the incubation were measured by ultraviolet spectrophotometer and circular dichroism spectrum, respectively.【Result】According to the results, the phosphorylation level of myoglobin was significantly lower (P＜0.05) in AP group than that in control group at 6 h, which indicating that alkaline phosphatase can catalyze the dephosphorylation of myoglobin in vitro, thus decrease the phosphorylation level of myoglobin. The relative content of oxymyoglobin in AP group was significantly higher than that in control group, and the relative content of metmyoglobin was significantly lower than that in control group after 2 h. In brief, the automatic oxidation rate of myoglobin was lower and the redox stability of myoglobin was higher in AP group than that in control group. However, no significant difference (P>0.05) was observed betweenAP group and control group, which means that pH value of the incubation system was not changed by adding alkaline phosphatase. The results showed that the secondary structure of myoglobin was mostly α-helix. From 0 min to 6 h incubation, the contents of α-helix and β-sheet of myoglobin were almost unchanged in AP group, while the α-helix content of myoglobin increased and β-sheet content of myoglobin decreased in control group, indicating that the secondary structural stability of Mb was increased after dephosphorylation.【Conclusion】It was speculated that the secondary structure of myoglobin might be changed after phosphorylation. The secondary structure stability was decreased and automatic oxidation rate of myoglobin was increased after phosphorylation. Thus, leading to the accumulation of metmyoglobin and color deterioration, and this might be one of the reasons by which protein phosphorylation play a negative role in regulating meat color stability.

Key words: meat color, myoglobin, phosphorylation, redox stability, secondary structural

LI Meng, LI Zheng, LI Xin, DU ManTing, SONG Xuan, ZHANG DeQuan . Effect of Phosphorylation Level on Myoglobin Stability[J].Scientia Agricultura Sinica, 2017, 50(22): 4382-4388.

0
/ / Recommend

Add to citation manager EndNote|Reference Manager|ProCite|BibTeX|RefWorks

URL: https://www.chinaagrisci.com/EN/10.3864/j.issn.0578-1752.2017.22.014

https://www.chinaagrisci.com/EN/Y2017/V50/I22/4382

References

[1] 尹靖东. 动物肌肉生物学与肉品科学. 北京: 中国农业大学出版社, 2011.

Yin J D. Animal Muscle Biology and Meat Quality. Beijing: China Agricultural University Press, 2011. ( in Chinese)

[2] Clydesdale F M. Colorimetry – methodology and applications. Critical Reviews in Food Science and Nutrition, 1978, 10: 243-301.

[3] Mancini R A, Hunt M C. Current research in meat color. Meat Science, 2005, 71(1): 100-121.

[4] Suman S P, Joseph P. Myoglobin chemistry and meat color. Annual Review of Food Science and Technology, 2013, 4: 79-99.

[5] 南庆贤. 肉类工业手册. 北京: 中国轻工业出版社, 2008.

Nan Q X. Meat Industry Handbook. Beijing: China Light Industry Press, 2008. ( in Chinese)

[6] 周光宏. 畜产品加工学. 北京: 中国农业出版社, 2011.

Zhou G H. Animal Products Processing. Beijing: China Agricultural Press, 2011. ( in Chinese)

[7] Canto A C, Suman S P, Nair M N, Li S, Rentfrow G, Beach C M, Silva T J, Wheeler T L, Shackelford S D, Grayson A, McKeith R O, King D A. Differential abundance of sarcoplasmic proteome explains animal effect on beef Longissimus lumborum color stability. Meat Science, 2015, 102: 90-98.

[8] Zhu K, Zhao J, Lubman D M, Miller F R, Barder T J. Protein pI Shifts due to Posttranslational Modifications in the Separation and Characterization of Proteins. Analytical Chemistry, 2005, 77(9): 2745-2755.

[9] Li M, Li X, Xin J, Li Z, Li G, Zhang Y, Du M, Shen Q, Zhang D. Effects of protein phosphorylation on color stability of ground meat. Food Chemistry, 2017, 219: 304-310.

[10] Tang J, Faustman C, Hoagland T A. Krzywicki revisited: equations for spectrophotometric determination of myoglobin redox forms in aqueous meat extracts. Journal of Food Science, 2004, 69(9): 717-720.

[11] Huang H, Larsen M R, Karlsson A H, Pomponio L, Costa L N, Lametsch R. Gel-based phosphoproteomics analysis of sarcoplasmic proteins in postmortem porcine muscle with pH decline rate and time differences. Proteomics, 2011, 11(20): 4063-4076.

[12] Huang H, Larsen M R, Lametsch R. Changes in phosphorylation of myofibrillar proteins during postmortem development of porcine muscle. Food Chemistry, 2012, 134(4): 1999-2006.

[13] Chen L, Li X, Ni N, Liu Y, Chen L, Wang Z, Shen Q, Zhang D. Phosphorylation of myofibrillar proteins in post-mortem ovine muscle with different tenderness. Journal of the Science of Food and Agriculture, 2016, 96(5): 1474-1483.

[14] Silverman-Gavrila L B, Lu T Z, Prashad R C, Nejatbakhsh N, Charlton M P, Feng Z P. Neural phosphoproteomics of a chronic hypoxia model-Lymnaea stagnalis. Neuroscience, 2009, 161(2): 621-634.

[15] Wang X, Ni M, Niu C, Zhu X, Zhao T, Zhu Z, Xuan Y, Cong W. Simple detection of phosphoproteins in SDS-PAGE by quercetin. Eupa open proteomics, 2014, 4: 156-164.

[16] Kendrew J C. Myoglobin and the structure of proteins. Science, 1963, 139: 1259-1266.

[17] Bekhit A E, Faustman C. Metmyoglobin reducing activity. Meat Science, 2005, 71(3): 407-39.

[18] Bisig D A, Di Iorio E E, Diederichs K, Winterhalter K H, Piontek K. Crystal structure of Asian elephant (Elephas maximus) cyano-metmyoglobin at 1.78-A resolution. Phe29 (B10) accounts for its unusual ligand binding properties.The Journal of Biological Chemistry, 1995, 270(35): 20754-20762.

[19] Suman S P, Faustman C, Stamer S L, Liebler D C. Proteomics of lipid oxidation-induced oxidation of porcine and bovine oxymyoglobins. Proteomics, 2007, 7(4): 628-640.

[20] Scheffler T L, Gerrard D E. Mechanisms controlling pork quality development: The biochemistry controlling postmortem energy metabolism. Meat Science, 2007, 77(1): 7-16.

[21] Gutzke D, Trout G R. Temperature and pH dependence of the autoxidation rate of bovine, ovine, porcine, and cervine oxymyoglobin isolated from three different muscless longissimus dorsi, gluteus medius, and biceps femoris. Journal of Agricultural and Food Chemistry, 2002, 50(9): 2673-2678.

[22] Shikama K, Sugawara Y. Autoxidation of native oxymyoglobin. Kinetic analysis of the pH profile. European Journal of Biochemistry, 1978, 91(2): 407-413.

[23] Ascenzi P, Marino M, Polticelli F, Coletta M, Gioia M, Marini S, Pesce A, Nardini M, Bolognesi M, Reeder B J, Wilson M T. Non-covalent and covalent modifications modulate the reactivity of monomeric mammalian globins. Biochimica et Biophysica Acta, 2013, 1834(9): 1750-1756.

[24] Schwägele F, Haschke C, Honikel K O, Krauss G. Enzymological investigations on the causes for the PSE-syndrome, I. Comparative studies on pyruvate kinase from PSE- and normal pig muscles. Meat Science, 1996, 44(1/2): 27-40.

[25] Sprang S R, Acharya K R, Goldsmith E J, Stuart D I, Varvill K, Fletterick R J, Madsen N B, Johnson L N. Structural changes in glycogen phosphorylase induced by phosphorylation. Nature, 1988, 336(6196): 215-221.

[26] Johnson L N. Glycogen phosphorylase: control by phosphorylation and allosteric effectors. FASEB Journal, 1992, 6(6): 2274-2282.

[27] Cai G Z, Callaci T P, Luther M A, Lee J C. Regulation of rabbit muscle phosphofructokinase by phosphorylation. Biophysical Chemistry, 1997, 64(1-3): 199-209.

[28] Kuo H J, Malencik D A, Liou R S, Anderson S R. Factors affecting the activation of rabbit muscle phosphofructokinase by actin. Biochemistry, 1986, 25(6): 1278-1286.

[29] Díaz-Moreno I, Hollingworth D, Frenkiel T A, Kelly G, Martin S, Howell S, García-Mayoral M, Gherzi R, Briata P, Ramos A. Phosphorylation-mediated unfolding of a KH domain regulates KSRP localization via 14-3-3 binding. Nature Structural & Molecular Biology, 2009, 16(3): 238-246.

[30] Zhang B, Liu J Y. Serine phosphorylation of the cotton cytosolic pyruvate kinase GhPK6 decreases its stability and activity. FEBS Open Biology, 2017, 7(3): 358-366.

Metrics

Viewed

Full text

Abstract

Cited

Shared

Discussed

Comments

Recommended 0

No Suggested Reading articles found!

Effect of Phosphorylation Level on Myoglobin Stability

RichHTML

PDF

Abstract

Cite this article

share this article

References

Related Articles 9

Metrics

Comments

Recommended 0

[1]	ZHANG YeJun,ZHANG DeQuan,HOU ChengLi,BAI YuQiang,REN Chi,WANG Xu,LI Xin. Effects of Protein Phosphorylation on the Dissociation and Acetylation Level of Actomyosin [J]. Scientia Agricultura Sinica, 2022, 55(7): 1433-1444.
[2]	YUAN Ke-jun, CHENG Lai-liang, NIU Qing-lin, WANG Jiang-yong. Identification and Analysis of Phosphoproteins in Red and Non-Red Apple Cultivars [J]. Scientia Agricultura Sinica, 2016, 49(8): 1530-1539.
[3]	CHEN Li-juan, LI Xin, LI Zheng, LI Pei-di, LI Zhong-wen, ZHANG De-quan. Protein Phosphorylation on the Function of Myofibrillar Proteins in Mutton Muscle [J]. Scientia Agricultura Sinica, 2016, 49(7): 1360-1370.
[4]	ZHANG Yan, LI Xin, LI Zheng, LI Meng, LIU Yong-feng, ZHANG De-quan. Effects of Controlled Freezing Point Storage on the Protein Phosphorylation Level [J]. Scientia Agricultura Sinica, 2016, 49(22): 4429-4440.
[5]	GAO Xing, LI Xin, LI Zheng, DU Man-ting, ZHANG Cai-xia, ZHANG De-quan, DING Wu. The Mechanism of Myosin Phosphorylation Regulating Actomyosin Dissociation of Skeletal Muscle During Postmortem [J]. Scientia Agricultura Sinica, 2016, 49(16): 3199-3207.
[6]	Lü Xiao-su, LI Yu-xuan, MIAO Ying, CHEN Liang-ke, SHEN Yuan-yue, QIN Ling, XING Yu. Analysis of Protein Phosphorylation Level at Different Developmental Stages of Strawberry Fruit [J]. Scientia Agricultura Sinica, 2016, 49(10): 1946-1959.
[7]	ZHOU Li-Sheng, YANG Jie, LIU Xian-Xian, ZHANG Zhi-Yan, YANG Bin, MA Jun-Wu. Genome-Wide Association Analyses for Musle pH 72 h Value and Meat Color Traits in Sutai Pigs [J]. Scientia Agricultura Sinica, 2014, 47(3): 564-573.
[8]	YE Zi, HUANG Cong-Cong, YU Rong. Microtubules in Guard Cells Involve in Serine/Threonine Phosphorylation During Stomatal Movement [J]. Scientia Agricultura Sinica, 2012, 45(21): 4351-4360.
[9]	GAO Yu-long,JU Xing-rong,QIU Wei-fen,WU Ding,JIANG Han-hu. Effects of High-Hydrostatic Pressure on the Membrane Damage and Oxidative Phosphorylation of Listeria monocytogenes [J]. Scientia Agricultura Sinica, 2011, 44(11): 2325-2333 .