role of n-terminal amino acids in the potency of anthrax lethal factor效力的n端氨基酸的炭疽致命因素.pdfVIP

Role of N-Terminal Amino Acids in the Potency of Anthrax Lethal Factor Pradeep K. Gupta, Mahtab Moayeri, Devorah Crown, Rasem J. Fattah, Stephen H. Leppla* Laboratory of Bacterial Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, United States of America Abstract Anthrax lethal factor (LF) is a Zn+2-dependent metalloprotease that cleaves several MAPK kinases and is responsible for the lethality of anthrax lethal toxin (LT). We observed that a recombinant LF (LF-HMA) which differs from wild type LF (LF-A) by the addition of two residues (His-Met) to the native Ala (A) terminus as a result of cloning manipulations has 3-fold lower potency toward cultured cells and experimental animals. We hypothesized that the ‘‘N-end rule’’, which relates the half-life of proteins in cells to the identity of their N-terminal residue, might be operative in the case of LF, so that the N-terminal residue of LF would determine the cytosolic stability and thereby the potency of LF. Mutational studies that replaced the native N-terminal residue of LF with known N-end rule stabilizing or destabilizing residues confirmed that the N-terminal residue plays a significant role in determining the potency of LT for cultured cells and experimental animals. The fact that a commercially-available LF preparation (LF-HMA) that is widely used in basic research studies and for evaluation of vaccines and therapeutics is 3-fold less potent than native LF (LF-A) should be considered when comparing published studies and in the design of future experiments. Citation: Gupta PK, Moayeri M, Crown D, Fattah RJ, Leppla SH (2008) Role of N-Terminal Amino Acids in the Potency of Anthrax Lethal Factor. PLoS ONE 3(9): e3130. doi:10.1371/journal.pone.0003130 Editor: Maxim Antopolsky, University of Helsinki, Finland Received July 2, 2008; Accepted August 12, 2008;