Structure-Function Relationship study of HupZ

NIH RePORTER · NIH · F31 · $40,174 · view on reporter.nih.gov ↗

Abstract

Abstract A heme utilization protein operon hupYZ encoding periplasmic proteins has recently been identified from Streptococcus pyogenes, a significant pathogen involved in a wide range of diseases. One of the encoded proteins, HupZ, was initially assigned as a heme oxygenase. However, our study revealed that this weak activity was due to a poly‐His‐tag induced structural similarity with the active site of genuine heme oxygenases. The tag‐free HupZ does not degrade heme; however, we recently discovered that tag‐free HupZ binds FMN. The FMN‐bound HupZ crystal structure superimposes with the biliverdin reductase (BVR) recently found in Mycobacterium tuberculosis (Mtb) that utilizes an F420H2 cofactor. The flavin moiety of FMN in HupZ aligns well with the deazaflavin moiety of the F420H2 cofactor. Therefore, we propose that HupZ is a flavin/deazaflavin oxidoreductase (FDOR) dependent BVR in the heme utilization pathway (Hup) of Streptococcus. Structural and functional characterizations are proposed to further the molecular understanding of this Hup protein using biochemical, spectroscopic, and structural approaches. We will interrogate how tag‐free HupZ interacts with its cofactor and substrate, and we will determine the critical residues involved in biliverdin reduction. We will attempt to characterize catalytic intermediates and elucidate the catalytic mechanism. Completing the proposed biochemical and structural studies will fill the gaps in our current understanding of the heme utilization pathway in a significant pathogen. Only a few FDOR‐dependent enzymes have been described, and the F420H2‐dependent BVR mechanism remains to be elucidated; therefore, this work is also highly significant to mechanistic enzymology.

Key facts

NIH application ID
10863866
Project number
5F31GM145187-03
Recipient
UNIVERSITY OF TEXAS SAN ANTONIO
Principal Investigator
Ephrahime Traore
Activity code
F31
Funding institute
NIH
Fiscal year
2024
Award amount
$40,174
Award type
5
Project period
2022-07-01 → 2025-06-30