Mb (myoglobin) plus H2O2 catalyses the oxidation of various substrates via a peroxidase-like activity. A Y103F (Tyr103→Phe) variant of human Mb has been constructed to assess the effect of exchanging an electron-rich oxidizable amino acid on the peroxidase activity of human Mb. Steady-state analyses of reaction mixtures containing Y103F Mb, purified linoleic acid and H2O2 revealed a lower total yield of lipid oxidation products than mixtures containing the wild-type protein, consistent with the reported decrease in the rate constant for reaction of Y103F Mb with H2O2 [Witting, Mauk and Lay (2002) Biochemistry 41, 11495–11503]. Irrespective of the Mb employed, lipid oxidation yielded 9(R/S)-HODE [9(R,S)-hydroxy-10E,12Z-octadecadienoic acid] in preference to 13(R/S)-HODE [13(R,S)-hydroxy-9Z,11E-octadecadienoic acid], while 9- and 13-keto-octadecadienoic acid were formed in trace amounts. However, lipid oxidation by the Y103F variant of Mb proceeded with a lower Vmax value and an increased Km value relative to the wild-type control. Consistent with the increased Km, the product distribution from reactions with Y103F Mb showed decreased selectivity compared with the wild-type protein, as judged by the decreased yield of 9(S)-relative to 9(R)-HODE. Together, these data verify that Tyr103 plays a significant role in substrate binding and orientation in the haem pocket of human Mb. Also, the midpoint potential for the Fe(III)/(II) one-electron reduction was shifted slightly, but significantly, to a higher potential, confirming the importance of Tyr103 to the hydrogen-bonding network involving residues that line the haem crevice of human Mb.
Skip Nav Destination
Close
Article navigation
July 2004
- Cover Image
- PDF Icon PDF LinkFront Matter
- PDF Icon PDF LinkTable of Contents
- PDF Icon PDF LinkEditorial Board
Research Article|
July 06 2004
Regio- and stereo-chemical oxidation of linoleic acid by human myoglobin and hydrogen peroxide: Tyr103 affects rate and product distribution
Benjamin S. RAYNER
;
Benjamin S. RAYNER
*Vascular Biology Group, ANZAC Research Institute, Hospital Road, Concord Repatriation General Hospital, Concord, NSW 2139, Australia
†Centre for Vascular Research, University of New South Wales, Sydney, NSW 2052, Australia
Search for other works by this author on:
Roland STOCKER
;
Roland STOCKER
†Centre for Vascular Research, University of New South Wales, Sydney, NSW 2052, Australia
Search for other works by this author on:
Peter A. LAY
;
Peter A. LAY
§Centre for Heavy Metal Research, School of Chemistry, University of Sydney, Sydney, NSW 2006, Australia
Search for other works by this author on:
Paul K. WITTING
Paul K. WITTING
1
*Vascular Biology Group, ANZAC Research Institute, Hospital Road, Concord Repatriation General Hospital, Concord, NSW 2139, Australia
†Centre for Vascular Research, University of New South Wales, Sydney, NSW 2052, Australia
1To whom correspondence should be addressed, at the ANZAC Research Institute (e-mail: pwitting@anzac.edu.au).
Search for other works by this author on:
Biochem J (2004) 381 (2): 365–372.
Article history
Received:
December 12 2003
Revision Received:
March 18 2004
Accepted:
March 22 2004
Accepted Manuscript online:
March 22 2004
Citation
Benjamin S. RAYNER, Roland STOCKER, Peter A. LAY, Paul K. WITTING; Regio- and stereo-chemical oxidation of linoleic acid by human myoglobin and hydrogen peroxide: Tyr103 affects rate and product distribution. Biochem J 15 July 2004; 381 (2): 365–372. doi: https://doi.org/10.1042/BJ20031924
Download citation file:
Close
Sign in
Don't already have an account? Register
Sign in to your personal account
You could not be signed in. Please check your email address / username and password and try again.
Biochemical Society Member Sign in
Sign InSign in via your Institution
Sign in via your InstitutionGet Access To This Article
Cited By
Related Articles
Engineering peroxidase activity in myoglobin: the haem cavity structure and peroxide activation in the T67R/S92D mutant and its derivative reconstituted with protohaemin-l-histidine
Biochem J (February,2004)
Tubular reabsorption rates for myoglobin in the isolated perfused rat kidney
Clin Sci (Lond) (June,1986)
Dynamics in transient complexes of redox proteins
Biochem Soc Trans (March,2012)