3 research outputs found
Moiety-Linkage Map Reveals Selective Nonbisphosphonate Inhibitors of Human Geranylgeranyl Diphosphate Synthase
Bisphosphonates
are potent inhibitors of farnesyl pyrophosphate synthase (FPPS) and
geranylgeranyl diphosphate synthase (GGPPS). Current bisphosphonate
drugs (e.g., Fosamax and Zometa) are highly efficacious in the treatment
of bone diseases such as osteoporosis, Paget’s disease, and
tumor-induced osteolysis, but they are often less potent in blood
and soft-tissue due to their phosphate moieties. The discovery of
nonbisphosphonate inhibitors of FPPS and/or GGPPS for the treatment
of bone diseases and cancers is, therefore, a current goal. Here,
we propose a moiety-linkage-based method, combining a site-moiety
map with chemical structure rules (CSRs), to discover nonbisphosphonate
inhibitors from thousands of commercially available compounds and
known crystal structures. Our moiety-linkage map reveals the binding
mechanisms and inhibitory efficacies of 51 human GGPPS (hGGPPS) inhibitors.
To the best of our knowledge, we are the first team to discover two
novel selective nonbisphosphonate inhibitors, which bind to the inhibitory
site of hGGPPS, using CSRs and site-moiety maps. These two compounds
can be considered as a novel lead for the potent inhibitors of hGGPPS
for the treatment of cancers and mevalonate-pathway diseases. Moreover,
based on our moiety-linkage map, we identified two key residues of
hGGPPS, K202, and K212, which play an important role for the inhibitory
effect of zoledronate (IC<sub>50</sub> = 3.4 ÎĽM and 2.4 ÎĽM,
respectively). This result suggests that our method can discover specific
hGGPPS inhibitors across multiple prenyltransferases. These results
show that the compounds that highly fit our moiety-linkage map often
inhibit hGGPPS activity and induce tumor cell apoptosis. We believe
that our method is useful for discovering potential inhibitors and
binding mechanisms for pharmaceutical targets
Control Activity of Yeast Geranylgeranyl Diphosphate Synthase from Dimer Interface through H‑Bonds and Hydrophobic Interaction
Previously we showed that
yeast geranylgeranyl diphosphate synthase (GGPPS) becomes an inactive
monomer when the first N-terminal helix involved in dimerization is
deleted. This raises questions regarding why dimerization is required
for GGPPS activity and which amino acids in the dimer interface are
essential for dimerization-mediated activity. According to the GGPPS
crystal structure, three amino acids (N101, N104, and Y105) located
in the helix F of one subunit are near the active site of the other
subunit. As presented here, when these residues were replaced individually
with Ala caused insignificant activity changes, N101A/Y105A and N101A/N104A
but not N104A/Y105A showed remarkably decreased <i>k</i><sub>cat</sub> values (200–250-fold). The triple mutant N101A/N104A/Y105A
displayed no detectable activity, although dimer was retained in these
mutants. Because N101 and Y105 form H-bonds with H139 and R140 in
the other subunit, respectively, we generated H139A/R140A double mutant
and found it was inactive and became monomeric. Therefore, the multiple
mutations apparently influence the integrity of the catalytic site
due to the missing H-bonding network. Moreover, Met111, also on the
highly conserved helix F, was necessary for dimer formation and enzyme
activity. When Met111 was replaced with Glu, the negative-charged
repulsion converted half of the dimer into a monomer. In conclusion,
the H-bonds mainly through N101 for maintaining substrate binding
stability and the hydrophobic interaction of M111 in dimer interface
are essential for activity of yeast GGPPS
Role of N‑Linked Glycans in the Interactions of Recombinant HCV Envelope Glycoproteins with Cellular Receptors
Hepatitis C virus (HCV) infection
is a major cause of chronic hepatitis
and hepatocellular carcinoma. It infects human liver cells through
several cellular protein receptors including CD81, SR-BI, claudin-1,
and occludin. Previous reports also show that lectin receptors can
mediate HCV recognition and entry. The envelope proteins of HCV (E1
and E2) are heavily glycosylated, further indicating the possible
roles of lectin receptor–virus interaction in HCV infection.
However, there is limited study investigating the relationship of
HCV envelope glycoproteins and lectin as well as non-lectin receptors.
Here we used surface plasmon resonance to examine the binding affinity
of different glycoforms of recombinant HCV envelope protein to receptors
and inspected the infectivity and assembly of HCV pseudoparticles
composed of different glycoforms of envelope proteins. Our results
indicated that DC-SIGN, L-SIGN, and Langerin had higher affinity to
recombinant HCV envelope proteins in the presence of calcium ions
than non-lectin receptors, and envelope proteins with Man8/9 N-glycans
showed approximate 10-fold better binding to lectin receptors than
envelope proteins with Man5 and complex type N-glycans. Interestingly,
comparing among glycoforms, recombinant envelope proteins with Man5
N-glycans showed the highest binding affinity when interacting with
non-lectin receptors. In summary, the glycans on HCV envelope protein
play a modulatory role in HCV assembly and infection and direct HCV–receptor
interaction, which mediates viral entry in different cells. Receptors
with high affinity to HCV envelope proteins may be considered as targets
for development of a therapeutic strategy against HCV