A Kinetic and Isotope Effect Investigation of the Urease-Catalyzed Hydrolysis of Hydroxyurea

作者：Marlier John F^*; Robins Lori I; Tucker Kathryn A; Rawlings Jill; Anderson Mark A; Cleland W W

来源：Biochemistry, 2010, 49(37): 8213-8219.

DOI：10.1021/bi100890v

The urease-catalyzed hydrolysis of hydroxyurea is known to exhibit biphasic kinetics, showing a rapid burst phase followed by a slow plateau phase. Kinetic isotope effects for both phases of this reaction were measured at pH 6.0 and 25 degrees C. The observed nitrogen isotope effects for the ammonia leaving group [(15)(V/K)NH(3)] were 1.0016 +/- 0.0005 during the burst phase and 1.0019 +/- 0.0007 during the plateau phase, while those for the hydroxylamine leaving group [(15)(V/K)NH(2)OH] were 1.0013 +/- 0.0005 for the burst phase and 1.0022 +/- 0.0003 for the plateau phase. These isotope effects are consistent with a rate-determining step that occurs prior to breaking either of the two possible C-N bonds. The observed carbonyl carbon isotope effects [(13)(V/K)] were 1.0135 +/- 0.0003 during the burst phase and 1.0178 +/- 0.0003 during the plateau phase. The similarity of the magnitude of the carbon isotope effects argues for formation of a common intermediate during both phases.

• 出版日期2010-9-21