5 research outputs found
Neutron Diffraction of Acetazolamide-Bound Human Carbonic Anhydrase II Reveals Atomic Details of Drug Binding
Carbonic anhydrases (CAs) catalyze the hydration of CO<sub>2</sub> forming HCO<sub>3</sub><sup>–</sup> and a proton,
an important
reaction for many physiological processes including respiration, fluid
secretion, and pH regulation. As such, CA isoforms are prominent clinical
targets for treating various diseases. The clinically used acetazolamide
(AZM) is a sulfonamide that binds with high affinity to human CA isoform
II (HCAÂ II). There are several X-ray structures available of
AZM bound to various CA isoforms, but these complexes do not show
the charged state of AZM or the hydrogen atom positions of the protein
and solvent. Neutron diffraction is a useful technique for directly
observing H atoms and the mapping of H-bonding networks that can greatly
contribute to rational drug design. To this end, the neutron structure
of H/D exchanged HCAÂ II crystals in complex with AZM was determined.
The structure reveals the molecular details of AZM binding and the
charged state of the bound drug. This represents the first determined
neutron structure of a clinically used drug bound to its target
Carbonic Anhydrase Inhibition with Benzenesulfonamides and Tetrafluorobenzenesulfonamides Obtained via Click Chemistry
A series
of novel benzene- and 2,3,5,6-tetrafluorobenzenesulfonamide was synthesized
by using a click chemistry approach starting from azido-substituted
sulfonamides and alkynes, incorporating aryl, alkyl, cycloalkyl, and
amino-/hydroxy-/halogenoalkyl moieties. The new compounds were medium
potency inhibitors of the cytosolic carbonic anhydrase (CA, EC 4.2.1.1)
isoforms I and II and low nanomolar/subnanomolar inhibitors of the
tumor-associated hCA IX and XII isoforms. The X-ray crystal structure
of two such sulfonamides in adduct with hCA II allowed us to understand
the factors governing inhibitory power
Dithiocarbamates Strongly Inhibit Carbonic Anhydrases and Show Antiglaucoma Action in Vivo
A series of dithiocarbamates were prepared by reaction
of primary/secondary amines with carbon disulfide in the presence
of bases. These compounds were tested for the inhibition of four human
(h) isoforms of the zinc enzyme carbonic anhydrase, CA (EC 4.2.1.1),
hCA I, II, IX, and XII, involved in pathologies such as glaucoma (CA
II and XII) or cancer (CA IX). Several low nanomolar inhibitors targeting
these CAs were detected. The X-ray crystal structure of the hCA II
adduct with morpholine dithiocarbamate evidenced the inhibition mechanism
of these compounds, which coordinate to the metal ion through a sulfur
atom from the dithiocarbamate zinc-binding function. Some dithiocarbamates
showed an effective intraocular pressure lowering activity in an animal
model of glucoma
Water Networks in Fast Proton Transfer during Catalysis by Human Carbonic Anhydrase II
Variants of human carbonic anhydrase II (HCA II) with
amino acid
replacements at residues in contact with water molecules in the active-site
cavity have provided insights into the proton transfer rates in this
protein environment. X-ray crystallography and <sup>18</sup>O exchange
measured by membrane inlet mass spectrometry have been used to investigate
structural and catalytic properties of variants of HCA II containing
replacements of Tyr7 with Phe (Y7F) and Asn67 with Gln (N67Q). The
rate constants for transfer of a proton from His64 to the zinc-bound
hydroxide during catalysis were 4 and 9 μs<sup>–1</sup> for Y7F and Y7F/N67Q, respectively, compared with a value of 0.8
μs<sup>–1</sup> for wild-type HCA II. These higher values
observed for Y7F and Y7F/N67Q HCA II could not be explained by differences
in the values of the p<i>K</i><sub>a</sub> of the proton
donor (His64) and acceptor (zinc-bound hydroxide) or by the orientation
of the side chain of the proton shuttle residue His64. They appeared
to be associated with a reduced level of branching in the networks
of hydrogen-bonded water molecules between proton shuttle residue
His64 and the zinc-bound solvent molecule as observed in crystal structures
at 1.5–1.6 Å resolution. Moreover, Y7F/N67Q HCA II is
unique among the variants studied in having a direct, hydrogen-bonded
chain of water molecules between the zinc-bound solvent and N<sup>ε</sup> of His64. This study provides the clearest example
to date of the relevance of ordered water structure to rate constants
for proton transfer in catalysis by carbonic anhydrase
Tricyclic Sulfonamides Incorporating Benzothiopyrano[4,3‑<i>c</i>]pyrazole and Pyridothiopyrano[4,3‑<i>c</i>]pyrazole Effectively Inhibit α- and β‑Carbonic Anhydrase: X‑ray Crystallography and Solution Investigations on 15 Isoforms
Carbonic anhydrases (CAs, EC 4.2.1.1) are ubiquitous
isozymes involved
in crucial physiological and pathological events, representing the
targets of inhibitors with several therapeutic applications. In this
connection, we report a new class of carbonic anhydrase inhibitors,
based on the thiopyrano-fused pyrazole scaffold to which a pendant
4-sulfamoylphenyl moiety was attached. The new sulfonamides <b>3a</b>–<b>e</b> were designed as constrained analogues
of celecoxib and valdecoxib. The most interesting feature of sulfonamides <b>3</b> was their predominantly strong inhibition of human (h) CA
I and II, as well as those of the mycobacterial β-class enzymes
(Rv1284, Rv3273, and Rv3588c), whereas their inhibitory action against
hCA III, IV, VA, VB, VI, VII, IX, XII, XIII, and XIV was found to
be at least 2 orders of magnitude lower. X-ray crystallography and
structural superposition studies made it possible to explain the very
distinct inhibition profile of the tricyclic sulfonamides, different
from those of celecoxib and valdecoxib