Affiliation:
1. Department of Microbiology & Immunology, Emory University, Atlanta, Georgia 30322
Abstract
ABSTRACT
The abilities of two bacterial active heme transporters, HmbR of
Neisseria meningitidis
and HemR of
Yersinia enterocolitica
, to use different heme sources were compared. While HmbR-expressing cells used only hemoglobin (Hb) and heme, HemR-expressing bacteria were able to grow on Hb, heme, myoglobin, hemopexin, catalase, human and bovine serum albumin-heme, and haptoglobin-hemoglobin complexes as sources of iron. Expression of functional HemR allowed
Escherichia coli
cells to respond to heme-containing peptides, microperoxidases MP-8, MP-9, and MP-11, suggesting the ability of HemR to transport heme covalently linked to other molecules. Comparison of HemR with other heme receptors identified several highly conserved histidine residues as well as two conserved amino acid motifs, the FRAP and NPNL boxes. A site-directed mutagenesis approach was used to investigate the roles of His128, His192, His352, and His461 residues in HemR function. The HemR receptor with histidine changed to lysine at position 128 (HemR
H128K
), HemR
H461L
, HemR
H461A
, and HemR
H128A,H461A
mutant receptors were unable to use Hb, human serum albumin-heme, and myoglobin as sources of porphyrin and iron. Utilization of free heme was also severely affected, with some residual heme uptake in cells expressing HemR
H128K
, HemR
H461A
, and HemR
H461L
. Conversely, the HemR
H192T
, HemR
H352A
, HemR
H352K
, and HemR
H192T,H352K
mutant receptors were fully functional. All mutant HemR proteins were expressed in the outer membrane at levels similar to that of the wild-type HemR receptor. Nonfunctional HemRs were able to bind heme- and Hb-agarose. A hypothetical model of the HemR function in which two conserved histidine residues, His128 and His461, participate in the transport of heme through the receptor pore is postulated.
Publisher
American Society for Microbiology
Subject
Molecular Biology,Microbiology
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