Identification of the Key Protein for Zinc Uptake in Hemophilus influenzae

Desheng Lu,Beth Boyd,Clifford A Lingwood

doi:10.1074/jbc.272.46.29033

Desheng Lu, Beth Boyd + Show 1 more

Open Access

https://doi.org/10.1074/jbc.272.46.29033

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Abstract

Very little is known about specific mechanisms for zinc accumulation and transport in bacteria. In this study a putative adhesin B in Hemophilus influenzae, the product of gene HI0119, has been identified as a periplasmic zinc-binding protein (PZP1). A pzp1-deficient mutant has been constructed which is defective for growth under aerobic conditions and grows poorly under anaerobic conditions. The growth defect is specifically rescued by supplementing the growth medium with high concentrations of zinc. Subcellular fractionation was used to localize PZP1 to the periplasmic region in a nontypeable H. influenzae strain and in a transfected recombinant Escherichia coli strain (TApzp1). Recombinant PZP1, purified from a periplasmic extract of E. coli strain TApzp1, contained approximately two zinc atoms/protein molecule as determined by neutron activation analysis and atomic absorption spectroscopy. The zinc atoms could be removed by incubation with EDTA, and, by further addition of zinc, a total of five zinc atoms/PZP1 could be bound. Direct binding of 65Zn to the recombinant protein by Western blot was demonstrated. Taken together, these results provide direct evidence that PZP1 plays a key role in zinc uptake by H. influenzae.

Highlights

Zinc is essential for all organisms because it plays a critical role in the catalytic activity and/or structural stability of many proteins
Direct binding of 65Zn to the recombinant protein by Western blot was demonstrated. These results provide direct evidence that PZP1 plays a key role in zinc uptake by H. influenzae
We demonstrate that this putative adhesin B is a periplasmic zinc-binding protein (PZP1) which plays a key role in the zinc uptake of H. influenzae

Summary

Introduction

Zinc is essential for all organisms because it plays a critical role in the catalytic activity and/or structural stability of many proteins. We reported the partial characterization of HI0119, identified from the H. influenzae genomic sequence [13] as a putative adhesin B because of its homology with the adhesin fimA of Streptococcus parasanguis This 37-kDa protein is distinct from fimA because of a central histidine-rich domain, potent celite binding ability, and a COOH-terminal disulfide-bonded domain. We demonstrate that this putative adhesin B is a periplasmic zinc-binding protein (PZP1) which plays a key role in the zinc uptake of H. influenzae. This is the first description of a potential prokaryotic zinc-specific transport protein

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