Date of this Version
Molecular Oral Microbiology
Aggregatibacter actinomycetemcomitans, a common inhabitant of the human upper aerodigestive tract, produces a repeat in toxin (RTX), leukotoxin (LtxA). The LtxA is transcribed as a 114-kDa inactive protoxin with activation being achieved by attachment of short chain fatty acyl groups to internal lysine residues. Methyl esters of LtxA that were isolated from A. actinomycetemcomitans strains JP2 and HK1651 and subjected to gas chromatography/mass spectrometry contained palmitoyl (C16:0, 27–29%) and palmitolyl (C16:1 cis Δ9, 43–44%) fatty acyl groups with smaller quantities of myristic (C14:0, 14%) and stearic (C18:0, 12–14%) fatty acids. Liquid chromatography/mass spectrometry of tryptic peptides from acylated and unacylated recombinant LtxA confirmed that Lys562 and Lys687 are the sites of acyl group attachment. During analysis of recombinant LtxA peptides, we observed peptide spectra that were not observed as part of the RTX acylation schemes of either Escherichia coli α-hemolysin or Bordetella pertussis cyclolysin. Mass calculations of these spectra suggested that LtxA was also modified by the addition of monohydroxylated forms of C14 and C16 acyl groups. Multiple reaction monitoring mass spectrometry identified hydroxymyristic and hydroxypalmitic acids in wild-type LtxA methyl esters. Single or tandem replacement of Lys562 and Lys687 with Arg blocks acylation, resulting in a >75% decrease in cytotoxicity when compared with wild-type toxin, suggesting that these posttranslational modifications are playing a critical role in LtxA-mediated target cell cytotoxicity.
This is the pre-peer reviewed version of the following article: [Fong, K. P., Tang, H.-Y., Brown, A. C., Kieba, I. R., Speicher, D. W., Boesze-Battaglia, K., & Lally, E. T. (2011). Aggregatibacter actinomycetemcomitans leukotoxin is post-translationally modified by addition of either saturated or hydroxylated fatty acyl chains. Molecular Oral Microbiology, 26(4), 262–276. http://doi.org/10.1111/j.2041-1014.2011.00617.x], which has been published in final form at [Link to final article using the http://doi.org/10.1111/j.2041-1014.2011.00617.x]. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions.
Aggregatibacter actinomycetemcomitans, RTX toxins, leukotoxin, acylation, post-translational protein modification, gas chromatography/mass spectrometry, reverse-phase liquid chromatography/tandem mass spectrometry
Fong, K. P., Tong, H. Y., Brown, A. C., Keiba, I. R., Speicher, D. W., Boeze-Battaglia, K., & Lally, E. T. (2011). Aggregatibacter Actinomycetemcomitans Leukotoxin is Post-Translationally Modified by Addition of Either Saturated or Hydroxylated Fatty Acyl Chains. Molecular Oral Microbiology, 26 (4), 262-276. http://dx.doi.org/10.1111/j.2041-1014.2011.00617.x
Date Posted: 01 March 2022
This document has been peer reviewed.