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Author Notes:
Correspondence: VA Medical Center Atlanta, Research Service 151, 1670 Clairmont Rd., Decatur, GA 30033, USA; Phone: +1 404 321-6111, ext. 3854; Fax: +1 404 329-2210; Email: dzahner@emory.edu
Acknowledgments: We thank Dr. Christian Klein (Bruker Daltonics) for his help with interpretation of ESI-MS/MS data of the crosslinks and Dr. Fred Strobel of the Department of Chemistry (Emory University) for exact mass determinations by ESI MS.
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Research Funding:
This work was supported by funds from NIH grant #R01AI 070829 (DSS) and a VA Merit Award from the Dept. of Veterans Affairs (DSS).
Keywords:
- fimbrial protein
- intramolecular cross-link
- proteolytic stability
- thermal stability
- mass spectrometry
Pilus backbone protein PitB of Streptococcus pneumoniae contains stabilizing intramolecular isopeptide bonds
Abstract:
Streptococcus pneumoniae type 2 pili are recently identified fimbrial structures extending from the bacterial surface and formed by polymers of the structural protein PitB. Intramolecular isopeptide bonds are a characteristic of the related pilus backbone protein Spy0128 of group A streptococci. Based on the identification of conserved residues in PitB, we predicted two intramolecular isopeptide bonds in PitB. Using a combination of tandem mass spectrometry and Edman sequencing, we show that these bonds were formed between Lys63-Asn214 and Lys243-Asn372 in PitB. Mutant proteins lacking the intramolecular isopeptide bonds retained the proteolytic stability observed with the wild type protein. However, absence of these bonds substantially decreased the melting temperature of the PitB-derivatives, indicating a stabilizing function of these bonds in PitB of the pneumococcal type 2 pilus.
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© 2011 Elsevier Inc. All rights reserved.
This is an Open Access work distributed under the terms of the Creative Commons Attribution-NonCommerical-NoDerivs 3.0 Unported License (http://creativecommons.org/licenses/by-nc-nd/3.0/).
