Characterization and kinetics studies of water buffalo (Bubalus bubalis) myoglobin.
The colour of buffalo (Bubalus bubalis L.) meat is darker than bovine meat. Since meat colour depends on the concentration of myoglobin (Mb) and its oxidation state, we have determined the main structural and functional properties of buffalo Mb. Buffalo Mb was purified from longissimus dorsi muscles and its molecular mass determined by ESI Q-TOF mass spectrometry. The molecular mass 17,034.50 was 86.20 Da higher than the bovine Mb. This was confirmed by analysing its primary structure, using a combined approach based on Edman degradation and MALDI-TOF mass spectrometry. Comparing the amino acid sequences of both Mbs, we found three amino acid differences out of 153 amino acid residues. One is a conservative substitution (D(bov)141E(buf)), and the other two (A(bov)19T(buf) and A(bov)117D(buf)) are nonconservative. These amino acid substitutions are unlikely to cause structural changes because they are located far from the heme binding pocket, as revealed by the 3D structure of buffalo Mb elaborated by homology modelling. Stability analyses show no difference with the bovine Mb for helix E and only minor differences in the stability values for helices A and G. Moreover, autoxidation rates of purified buffalo and bovine myoglobins at 37 degrees C, pH 7.2, were almost identical, 0.052+/-0.001 h(-1) and 0.054+/-0.002 h(-1), respectively, as were their oxygen-binding Kd values, 3.7+/-0.1 microM and 3.5+/-0.1 microM, respectively. The percent of MetMb values were almost identical. The results presented here suggest that the darker buffalo meat depends on factors other than the oxidation rate of its Mb, as, for example, the Mb content (0.393+/-0.005 g/100 g of tissue) and consequently MetMb, which are almost twice as high as bovine meat (Mb: 0.209+/-0.003 g/100 g of tissue).
Dosi
Roberta
R
Department of Life Sciences, Second University of Naples, Via Vivaldi 43, I-81100 Caserta, Italy.
Di Maro
Antimo
A
Chambery
Angela
A
Colonna
Giovanni
G
Costantini
Susan
S
Geraci
Giuseppe
G
Parente
Augusto
A
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D002020:Buffaloes / Q000378:metabolism*; D002417:Cattle; D007700:Kinetics; D013058:Mass Spectrometry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009211:Myoglobin / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D010084:Oxidation-Reduction; D016415:Sequence Alignment
D009211:Myoglobin