First Advisor

Dirk Iwata-Reuyl

Date of Award

2016

Document Type

Thesis

Degree Name

Bachelor of Science (B.S.) in Biochemistry and University Honors

Department

Chemistry

Subjects

Enzymes, Coenzymes -- Synthesis, NAD (Coenzyme)

DOI

10.15760/honors.284

Abstract

The NADPH-dependent nitrile-oxidioreductase, QueF is the only known enzyme capable of reducing a nitrile group to an amine. This ability makes it an attractive alternative to conventional industrial nitrile reduction. Understating how QueF binds NADPH may lead to the development of an enzyme capable of using the less expensive reductive cofactor NADH. A cofactor docking model indicates that key residues, Q60 and Y21, interact with the ribose phosphate moiety unique to NADPH. Mutants of these residues (Q60A, Q60N, Q60E, Y21A and Y21F) were developed for steady-state kinetic analysis. Modification to either residue resulted in a decrease in binding and catalytic activity when using NADPH as a hydride source. Q60E had the greatest reduction in activity (0.45% of WT). There appears to be both charge and steric factors involved in direct cofactor binding. Neither WT or mutant enzymes were able to utilize NADH as a reductive cofactor to any observable extent.

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Persistent Identifier

http://archives.pdx.edu/ds/psu/17913

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