Visual Analysis and Inhibitor Screening of Leucine Aminopeptidase, a Key Virulence Factor for Pathogenic Bacteria-Associated Infection

Many pathogenic bacteria produce pore-forming toxins to attack and kill human cells. We have determined the 4.5 Å structure of the ~2.2 MDa pore complex of pneumolysin, the main virulence factor of Streptococcus pneumoniae, by cryoEM. The pneumolysin pore is a 400 Å ring of 42 membrane-inserted monomers. Domain 3 of the soluble toxin refolds into two ~85 Å β-hairpins that traverse the lipid bilayer and assemble into a 168-strand β-barrel. The pore complex is stabilized by salt bridges between β-hairpins of adjacent subunits and an internal α-barrel. The apolar outer barrel surface with large sidechains is immersed in the lipid bilayer, while the inner barrel surface is highly charged. Comparison of the cryoEM pore complex to the prepore structure obtained by electron cryo-tomography and the x-ray structure of the soluble form reveals the detailed mechanisms by which the toxin monomers insert into the lipid bilayer to perforate the target membrane.

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Reduced activity of the hypertension-associated Lys528Arg mutant of human adipocyte-derived leucine aminopeptidase (A-LAP)/ER-aminopeptidase-1

FEBS Letters ◽

10.1016/j.febslet.2006.02.041 ◽

2006 ◽

Vol 580 (7) ◽

pp. 1833-1838 ◽

Cited By ~ 76

Author(s):

Yoshikuni Goto ◽

Akira Hattori ◽

Yasuhiro Ishii ◽

Masafumi Tsujimoto

Keyword(s):

Leucine Aminopeptidase ◽

Human Adipocyte ◽

Aminopeptidase A ◽

Reduced Activity

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Microarray Analysis of Tomato’s Early and Late Wound Response Reveals New Regulatory Targets for Leucine Aminopeptidase A

PLoS ONE ◽

10.1371/journal.pone.0077889 ◽

2013 ◽

Vol 8 (10) ◽

pp. e77889 ◽

Cited By ~ 15

Author(s):

Melissa A. Scranton ◽

Jonathan H. Fowler ◽

Thomas Girke ◽

Linda L. Walling

Keyword(s):

Microarray Analysis ◽

Leucine Aminopeptidase ◽

Wound Response ◽

Aminopeptidase A

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Characterization of a leucine aminopeptidase of Babesia gibsoni

Parasitology ◽

10.1017/s0031182009006398 ◽

2009 ◽

Vol 136 (9) ◽

pp. 945-952 ◽

Cited By ~ 9

Author(s):

H. JIA ◽

M. A. TERKAWI ◽

G. O. ABOGE ◽

Y.-K. GOO ◽

Y. LUO ◽

...

Keyword(s):

Amino Acid ◽

Free Amino Acid ◽

Free Amino ◽

Drug Targets ◽

Leucine Aminopeptidase ◽

Divalent Cations ◽

Specific Inhibitor ◽

Babesia Gibsoni ◽

Aminopeptidase A

SUMMARYPeptidases of parasitic protozoa are currently under intense investigation in order to identify novel virulence factors, drug targets, and vaccine candidates, except in Babesia. Leucine aminopeptidases in protozoa, such as Plasmodium and Leishmania, have been identified to be involved in free amino acid regulation. We report here the molecular and enzymatic characterization, as well as the localization of a leucine aminopeptidase, a member of the M17 cytosolic aminopeptidase family, from B. gibsoni (BgLAP). A functional recombinant BgLAP (rBgLAP) expressed in Escherichia coli efficiently hydrolysed synthetic substrates for aminopeptidase, a leucine substrate. Enzyme activity of the rBgLAP was found to be optimum at pH 8·0 and at 37°C. The substrate profile was slightly different from its homologue in P. falciprum. The activity was also strongly dependent on metal divalent cations, and was inhibited by bestatin, which is a specific inhibitor for metalloprotease. These results indicated that BgLAP played an important role in free amino acid regulation.

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