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J Inorg Biochem. 2017 Feb;167:150-156. doi: 10.1016/j.jinorgbio.2016.08.015. Epub 2016 Aug 27.

Crucial residue Trp158 of lipoprotein PiaA stabilizes the ferrichrome-PiaA complex in Streptococcus pneumoniae.

Author information

1
Key Laboratory of Functional Protein Research of Guangdong Higher Education Institutes, Institute of Life and Health Engineering, College of Life Science and Technology, Jinan University, Guangzhou 510632, China.
2
Key Laboratory of Functional Protein Research of Guangdong Higher Education Institutes, Institute of Life and Health Engineering, College of Life Science and Technology, Jinan University, Guangzhou 510632, China. Electronic address: tsunxs@jnu.edu.cn.
3
Key Laboratory of Functional Protein Research of Guangdong Higher Education Institutes, Institute of Life and Health Engineering, College of Life Science and Technology, Jinan University, Guangzhou 510632, China. Electronic address: tqyhe@jnu.edu.cn.

Abstract

The pathogenic Streptococcus pneumoniae (S. pneumoniae) has evolved a special mechanism such as pneumococcal iron acquisition ATP binding cassette (PiaABC) to take up siderophore-iron from its host. The cell-surface lipoprotein PiaA, a key component of PiaABC, is the primary receptor to bind ferrichrome (Fc). To study the structure-function relationship of PiaA, three conservative amino-acid residues, Trp63, Trp158 and Phe255, in the hydrophobic barrel of the metal binding site of PiaA, were individually and collectively mutated to alanine; and the resulted single-point mutants, W63A, W158A and F255A, and triple mutant W63A/W158A/F255A were characterized by using biochemical and biophysical methods. Experiments showed that wild-type PiaA (WT-PiaA) and the single-point mutant proteins bound Fc with a similar kinetics mode, but the reaction rate of W158A was lower than that for WT-PiaA. The binding affinity of W158A toward Fc was significantly weaker than that of the WT-PiaA-Fc (wild-type PiaA bound with Fc) interaction. Furthermore, the absence of Trp158 in the protein led to a significant impact on the secondary structure of PiaA, resulting in a labile conformational structure of W158A, with impaired resistance to thermal and chemical denaturation. Collectively, Trp158 is a crucial residue for binding Fc, playing an important role in stabilizing the PiaA-Fc complex. This study revealed the critical role of the conserved tryptophan residues in Fc-binding protein PiaA, and provided valuable information for understanding the Fc transport mechanism mediated by PiaA or its homologous proteins in bacteria.

KEYWORDS:

Ferrichrome; Iron transport; Iron-binding protein; PiaA; Streptococcus pneumoniae

PMID:
28341101
DOI:
10.1016/j.jinorgbio.2016.08.015
[Indexed for MEDLINE]

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