EXPRESSION AND CHARACTERIZATION OF VMPH REVEALS NEW INSIGHTS INTO DIHEME ENZYMES

Cowart, Nikhil

Permalink

https://hdl.handle.net/2142/133274 Copy

Description

Title

EXPRESSION AND CHARACTERIZATION OF VMPH REVEALS NEW INSIGHTS INTO DIHEME ENZYMES

Author(s)

Cowart, Nikhil

Issue Date

2026-05-13

Director of Research (if dissertation) or Advisor (if thesis)

Manesis, Anastasia

Department of Study

Chemistry

Degree Granting Institution

University of Illinois at Urbana-Champaign

Degree Name

B.S. (bachelor's)

Degree Level

Thesis

Date of Ingest

2026-05-14T14:27:19-05:00

Keyword(s)

Diheme Cytochrome c Peroxidase, Copper Binding, MbnH, MbnP

Language

eng

Abstract

A recently emerging set of bacterial cytochrome c peroxidases (bCcPs), known as bisFe(IV) forming enzymes, form a high valent electronic intermediate that is different from canonical bCcPs. Bioinformatic analysis of the diheme peroxidase superfamily shows a number of larger enzymes, with a domain similar to bis-Fe(IV) forming enzymes, potentially “fused” with their putative partners. One of these enzymes, VMPH, is predicted to fold similarly to a bisFe(IV) forming enzyme called MbnH and its substrate MbnP. Expression optimization efforts of VMPH have revealed the importance of preventing excessive growth of the started culture, and an expression temperature of 20 C. Adhering to these methods allowed for the reproducible expression of soluble VMPH with around ~70% heme loading. Ongoing structural characterization efforts have shown progress in obtaining protein crystals needed to determine the structure of VMPH. These efforts have revealed the ability of VMPH to play a role in copper homeostasis. EPR and copper binding assays of VMPH reveal the copper binding affinity of VMPH as well as the ability to bind both Cu(I) and Cu(II). The log(kd) value of -16.150.40 falls well within the range of other copper binding proteins showing that VMPH could play a role in copper homeostasis.

Type of Resource

text

Owning Collections