Engineering Ascorbate Peroxidase Activity Into Cytochrome C Peroxidase

Journal Article · Tue May 26 04:00:00 EDT 2009 · Biochem.47:10324,2008

OSTI ID:953615

Meharenna, Y T; Oertel, P; Bhaskar, B; Poulos, T L

Cytochrome c peroxidase (CCP) and ascorbate peroxidase (APX) have very similar structures, and yet neither CCP nor APX exhibits each others activities with respect to reducing substrates. APX has a unique substrate binding site near the heme propionates where ascorbate H-bonds with a surface Arg and one heme propionate (Sharp et al. (2003) Nat. Struct. Biol. 10, 303--307). The corresponding region in CCP has a much longer surface loop, and the critical Arg residue that is required for ascorbate binding in APX is Asn in CCP. In order to convert CCP into an APX, the ascorbate-binding loop and critical arginine were engineered into CCP to give the CCP2APX mutant. The mutant crystal structure shows that the engineered site is nearly identical to that found in APX. While wild-type CCP shows no APX activity, CCP2APX catalyzes the peroxidation of ascorbate at a rate of {approx}12 min{sup -1}, indicating that the engineered ascorbate-binding loop can bind ascorbate.

• Research Organization: Stanford Linear Accelerator Center (SLAC)
• Sponsoring Organization: USDOE
• DOE Contract Number: AC02-76SF00515
• OSTI ID: 953615
• Report Number(s): SLAC-REPRINT-2009-268
• Journal Information: Biochem.47:10324,2008, Journal Name: Biochem.47:10324,2008 Journal Issue: 39 Vol. 47; ISSN 0006-2960; ISSN BICHAW
• Country of Publication: United States
• Language: English

Related Subjects

36 MATERIALS SCIENCE
ARGININE
CRYSTAL STRUCTURE
CYTOCHROMES
ENGINEERING
HEME
MUTANTS
Other
BIO
CHEM
PEROXIDASES
RESIDUES
SUBSTRATES
SURFACES