- Original Article
- Biochemistry/Molecular Biology
Purification and characterization of protocatechuate 3,4-dioxygenase from Pseudomonas pseudoalcaligenes KF707
Journal of the Korean Society for Applied Biological Chemistry volume 56, pages 401–408 (2013)
Protocatechuate 3,4-dioxygenase was isolated and characterized from Pseudomonas pseudoalcaligenes KF707 for the purpose of developing a new anti-browning agent. The protocatechuate 3,4-dioxygenase from Pseudomonas pseudoalcaligenes KF707 was purified 296.8-fold, and showed specific activity of 121.7 U/mg. Based on the SDS-polyacrylamide and gel permeation chromatography, the molecular weight of protocatechuate 3,4-dioxygenase was 189.9 kDa and was composed of 3 αβ protomers, with molecular weights of 29.0 kDa of α subunit and 34.3 kDa of â subunit. The optimal pH and temperature were 7.5 and 38°C, respectively. Km values of catechol, protocatechuate, gallate, p-cresol, caffeic acid, catechin, L-DOPA, 4-methylcatechol and pyrogallol were 14, 17, 2, 10, 12, 20, 30, 21 and 3 μM, and the Vmax/Km (mim−1) values were 0.052, 3.06, 0.35, 0.01, 0.03, 0.02, 0.006, 0.008 and 0.11, respectively. This indicates that the enzyme is active on a wide range of phenyl compounds, in contrast to the high specificity of similar enzymes from other sources. Our data also show that the turnover number of protocatechuate 3,4-dioxygenase from Pseudomonas pseudoalcaligenes KF707 is 68 s−1, which is much higher than the known values from other sources.
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H. W. Sim and M. J. Jung contributed equally.
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Sim, H.W., Jung, M. & Cho, Y.K. Purification and characterization of protocatechuate 3,4-dioxygenase from Pseudomonas pseudoalcaligenes KF707. J Korean Soc Appl Biol Chem 56, 401–408 (2013). https://doi.org/10.1007/s13765-013-3080-2