8 research outputs found
Structural Basis of Single-Nucleotide Polymorphisms in Cytochrome P450 2C9
Single-nucleotide
polymorphisms in drug-metabolizing cytochrome
P450 (CYP) enzymes are important contributors to interindividual differences
in drug metabolism leading to adverse drug reactions. Despite their
extensive characterization and importance in pharmacogenetics of clinical
drugs, the structural basis of CYP polymorphisms has remained scant.
Here we report the crystal structures of human CYP2C9 and its polymorphic
variants, *3 (I359L) and *30 (A477T), with an antihypertensive drug
losartan. The structures show distinct interaction and occupation
of losartan in the active site, the access channel, and the peripheral
binding site. The I359L substitution located far from the active site
remarkably altered the residue side chains near the active site and
the access channel, whereas the T477 substitution illustrated hydrogen-bonding
interaction with the reoriented side chain of Q214. The results yield
structural insights into the reduced catalytic activity of the CYP2C9
variants and have important implications for understanding genetic
polymorphisms in CYP-mediated drug metabolism
Structure Analysis and Derivation of Deformed Electron Density Distribution of Polydiacetylene Giant Single Crystal by the Combination of X‑ray and Neutron Diffraction Data
The
crystal structure of polydiacetylene giant single crystal has
been analyzed on the basis of the two different methods of wide-angle
neutron diffraction and X-ray diffraction. The X-ray result gives
us the total electron density distribution [<b>ρ</b>(<b>x</b>)] of polymer chain. The neutron result tells the positions
of atomic nuclei, which can allow us to speculate the electron density
distributions [<b>ρ</b><sub>0</sub>(<b>x</b>)] around
the nonbonded isolated atoms. As a result, the so-called bonded (or
deformed) electron density Δρ(<b>x</b>) [≡
ρ(<b>x</b>) – ρ<sub>0</sub>(<b>x</b>) = ρ<sub>X</sub>(<b>x</b>) – ρ<sub>N</sub>(<b>x</b>)], i.e., the electron density distribution due to
the conjugation among the covalently bonded atoms along the polymer
chain, can be estimated using the two information obtained by the
X-ray and neutron data analyses (the so-called X-ray–neutron
subtraction (X<i>–</i>N) method). The present report
is the first example of the application of X–N method to the
synthetic polymer species. The Δρ(<b>x</b>) derived
for polydiacetylene was found similar to that of the low-molecular-weight
model compound having the similar electronically conjugated chemical
formula. The Δρ(<b>x</b>) was calculated by the
density functional theory, which was in a good agreement with the
experimental result qualitatively
Structure Analysis and Derivation of Deformed Electron Density Distribution of Polydiacetylene Giant Single Crystal by the Combination of X‑ray and Neutron Diffraction Data
The
crystal structure of polydiacetylene giant single crystal has
been analyzed on the basis of the two different methods of wide-angle
neutron diffraction and X-ray diffraction. The X-ray result gives
us the total electron density distribution [<b>ρ</b>(<b>x</b>)] of polymer chain. The neutron result tells the positions
of atomic nuclei, which can allow us to speculate the electron density
distributions [<b>ρ</b><sub>0</sub>(<b>x</b>)] around
the nonbonded isolated atoms. As a result, the so-called bonded (or
deformed) electron density Δρ(<b>x</b>) [≡
ρ(<b>x</b>) – ρ<sub>0</sub>(<b>x</b>) = ρ<sub>X</sub>(<b>x</b>) – ρ<sub>N</sub>(<b>x</b>)], i.e., the electron density distribution due to
the conjugation among the covalently bonded atoms along the polymer
chain, can be estimated using the two information obtained by the
X-ray and neutron data analyses (the so-called X-ray–neutron
subtraction (X<i>–</i>N) method). The present report
is the first example of the application of X–N method to the
synthetic polymer species. The Δρ(<b>x</b>) derived
for polydiacetylene was found similar to that of the low-molecular-weight
model compound having the similar electronically conjugated chemical
formula. The Δρ(<b>x</b>) was calculated by the
density functional theory, which was in a good agreement with the
experimental result qualitatively
Structure Analysis and Derivation of Deformed Electron Density Distribution of Polydiacetylene Giant Single Crystal by the Combination of X‑ray and Neutron Diffraction Data
The
crystal structure of polydiacetylene giant single crystal has
been analyzed on the basis of the two different methods of wide-angle
neutron diffraction and X-ray diffraction. The X-ray result gives
us the total electron density distribution [<b>ρ</b>(<b>x</b>)] of polymer chain. The neutron result tells the positions
of atomic nuclei, which can allow us to speculate the electron density
distributions [<b>ρ</b><sub>0</sub>(<b>x</b>)] around
the nonbonded isolated atoms. As a result, the so-called bonded (or
deformed) electron density Δρ(<b>x</b>) [≡
ρ(<b>x</b>) – ρ<sub>0</sub>(<b>x</b>) = ρ<sub>X</sub>(<b>x</b>) – ρ<sub>N</sub>(<b>x</b>)], i.e., the electron density distribution due to
the conjugation among the covalently bonded atoms along the polymer
chain, can be estimated using the two information obtained by the
X-ray and neutron data analyses (the so-called X-ray–neutron
subtraction (X<i>–</i>N) method). The present report
is the first example of the application of X–N method to the
synthetic polymer species. The Δρ(<b>x</b>) derived
for polydiacetylene was found similar to that of the low-molecular-weight
model compound having the similar electronically conjugated chemical
formula. The Δρ(<b>x</b>) was calculated by the
density functional theory, which was in a good agreement with the
experimental result qualitatively
Identification of Highly Potent Human Immunodeficiency Virus Type‑1 Protease Inhibitors against Lopinavir and Darunavir Resistant Viruses from Allophenylnorstatine-Based Peptidomimetics with P2 Tetrahydrofuranylglycine
The emergence of
drug-resistant HIV from a widespread antiviral
chemotherapy targeting HIV protease in the past decades is unavoidable
and provides a challenge to develop alternative inhibitors. We synthesized
a series of allophenylnorstatine-based peptidomimetics with various
P<sub>3</sub>, P<sub>2</sub>, and P<sub>2</sub>́ moieties. The
derivatives with P<sub>2</sub> tetrahydrofuranylglycine (Thfg) were
found to be potent against wild type HIV-1 protease and the virus,
leading to a highly potent compound <b>21f</b> (KNI-1657) against
lopinavir/ritonavir- or darunavir-resistant strains. Co-crystal structures
of <b>21f</b> and the wild-type protease revealed numerous key
hydrogen bonding interactions with Thfg. These results suggest that
the strategy to design allophenylnorstatine-based peptidomimetics
combined with Thfg residue would be promising for generating candidates
to overcome multidrug resistance
Seven Cysteine-Deficient Mutants Depict the Interplay between Thermal and Chemical Stabilities of Individual Cysteine Residues in Mitogen-Activated Protein Kinase c‑Jun N‑Terminal Kinase 1
Intracellular proteins can have free cysteines that may
contribute to their structure, function, and stability; however, free
cysteines can lead to chemical instabilities in solution because of
oxidation-driven aggregation. The MAP kinase, c-Jun N-terminal kinase
1 (JNK1), possesses seven free cysteines and is an important drug
target for autoimmune diseases, cancers, and apoptosis-related diseases.
To characterize the role of cysteine residues in the structure, function,
and stability of JNK1, we prepared and evaluated wild-type JNK1 and
seven cysteine-deficient JNK1 proteins. The nonreduced sodium dodecyl
sulfate–polyacrylamide gel electrophoresis experiments showed
that the chemical stability of JNK1 increased as the number of cysteines
decreased. The contribution of each cysteine residue to biological
function and thermal stability was highly susceptible to the environment
surrounding the particular cysteine mutation. The mutations of solvent-exposed
cysteine to serine did not influence biological function and increased
the thermal stability. The mutation of the accessible cysteine involved
in the hydrophobic pocket did not affect biological function, although
a moderate thermal destabilization was observed. Cysteines in the
loosely
assembled hydrophobic environment moderately contributed to thermal
stability, and the mutations of these cysteines had a negligible effect
on enzyme activity. The other cysteines are involved in the tightly
filled hydrophobic core, and mutation of these residues was found
to correlate with thermal stability and enzyme activity. These findings
about the role of cysteine residues should allow us to obtain a stable
JNK1 and thus promote the discovery of potent JNK1 inhibitors
Identification of Highly Potent Human Immunodeficiency Virus Type‑1 Protease Inhibitors against Lopinavir and Darunavir Resistant Viruses from Allophenylnorstatine-Based Peptidomimetics with P2 Tetrahydrofuranylglycine
The emergence of
drug-resistant HIV from a widespread antiviral
chemotherapy targeting HIV protease in the past decades is unavoidable
and provides a challenge to develop alternative inhibitors. We synthesized
a series of allophenylnorstatine-based peptidomimetics with various
P<sub>3</sub>, P<sub>2</sub>, and P<sub>2</sub>́ moieties. The
derivatives with P<sub>2</sub> tetrahydrofuranylglycine (Thfg) were
found to be potent against wild type HIV-1 protease and the virus,
leading to a highly potent compound <b>21f</b> (KNI-1657) against
lopinavir/ritonavir- or darunavir-resistant strains. Co-crystal structures
of <b>21f</b> and the wild-type protease revealed numerous key
hydrogen bonding interactions with Thfg. These results suggest that
the strategy to design allophenylnorstatine-based peptidomimetics
combined with Thfg residue would be promising for generating candidates
to overcome multidrug resistance
Insights into the Proton Transfer Mechanism of a Bilin Reductase PcyA Following Neutron Crystallography
Phycocyanobilin, a light-harvesting
and photoreceptor pigment in
higher plants, algae, and cyanobacteria, is synthesized from biliverdin
IXα (BV) by phycocyanobilin:ferredoxin oxidoreductase (PcyA)
via two steps of two-proton-coupled two-electron reduction. We determined
the neutron structure of PcyA from cyanobacteria complexed with BV,
revealing the exact location of the hydrogen atoms involved in catalysis.
Notably, approximately half of the BV bound to PcyA was BVH<sup>+</sup>, a state in which all four pyrrole nitrogen atoms were protonated.
The protonation states of BV complemented the protonation of adjacent
Asp105. The “axial” water molecule that interacts with
the neutral pyrrole nitrogen of the A-ring was identified. His88 Nδ
was protonated to form a hydrogen bond with the lactam O atom of the
BV A-ring. His88 and His74 were linked by hydrogen bonds via H<sub>3</sub>O<sup>+</sup>. These results imply that Asp105, His88, and
the axial water molecule contribute to proton transfer during PcyA
catalysis