Inhibition of mammalian carbonic anhydrases I-XIV with grayanotoxin III: solution and in silico studies

Figures & data

Scheme 1. 2D molecular structures of GTX3, DZA, BRZ, DCP, and AAZ.

Table 1. Inhibition data of mammalian isoforms hCA I-XIV with grayanotoxin III, and the sulfonamide inhibitor dichlorophenamide DCP (as standard) by a stopped-flow CO₂ hydrase assay24.

	KI (µM)*
Compound/Isoform	GTX3	DCP
hCA I	8.01	1.20
hCA II	6.13	0.038
hCA III	0.76	680
hCA IV	0.51	15.0
hCA VA	1.14	0.63
hCA VB	0.74	0.021
hCA VI	0.86	0.079
hCA VII	1.26	0.026
hCA IX	0.98	0.050
hCA XII	2.15	0.050
hCA XIII	0.69	0.023
hCA XIV	1.47	0.34

*Mean from three different determinations. Errors were in the range of ± 10% of the reported values.

Figure 1. Binding of grayanotoxin III (GTX3) to hCA I (A) and hCA II (B). The protein backbone is represented with residue position default colors in Maestro, the catalytic Zn(II) ion as the pale brown sphere and GTX3 is shown as stick model (carbon gray, oxygen red, hydrogen white).

Figure 2. Detailed interactions of grayanotoxin III (GTX3) when bound to the hCA I active site. Docking score at hCA-I: −8.21 kcal/mol (Glide Induced Fit Docking). The protein backbone is represented with residue position default colors in Maestro, the catalytic Zn(II) ion as the pale brown sphere and GTX3 is shown as stick model (carbon gray, oxygen red, hydrogen white). Amino acid residues and water molecules involved in the interaction with grayanotoxin III are also evidenced (CPK colors; hCA I numbering system).

Figure 3. Detailed interactions of grayanotoxin III (GTX3) when bound to the hCA II active site. Docking score at CA-II: −8.29 kcal/mol; (Glide Induced Fit Docking). The protein backbone is represented with residue position default colors in Maestro, the catalytic Zn(II) ion as the pale brown sphere and GTX3 is shown as stick model (carbon gray, oxygen red, hydrogen white). Amino acid residues and water molecules involved in the interaction with grayanotoxin III are also evidenced (CPK colors; hCA I numbering system).

Figure 4. Ligand interaction diagrams for (A) hCA I and (B) hCA II. Residues are labeled and colored according to residue type (green, hydrophobic; cyan, polar; red, negative; purple, positive). Yellow markers on the ligand denote per-atom solvent accessible surface area and marker size shows the amount of exposure. Labeling of residue (font sizes) is used to represent the depth of a residue in the three dimensional view (i.e. “closer” to the viewer are shown in a large font, whereas, residues that are “far away” are shown with small font. While hydrogen bonds to side chains are represented with dashed, hydrogen bonds to backbones are represented as solid lines.

Table 2. Details of molecular docking results of GTX3 within the active sites of hCA I and hCA II using Induced Fit Docking (Glide/XP). Energy units are in kcal/mol.

	Glide_ Gscore	Glide_Lig_ efficiency In	Glide_ energy	Glide_ hbond	Glide_ lipo	Glide_ Ecoul	Glide_ Einter	Glide_ Evdw	Glide_Lig_ Efficiency	Glide_Lig_ eff_SA	Prime_ Coulomb	Prime_ Covalent	Prime_ VDW	Prime_ Sol_SA	Prime_ Sol_GB	Prime_ Energy	XP_ Hbond	IFD_ Score
hCA-1	−8.21	−1.93	−39.54	−0.91	−1.50	−13.81	0.91	−25.73	−0.32	−0.94	−8801.40	1346.85	−1325.02	59.50	−1007.33	−9727.40	−2.82	−494.58
hCA-2	−8.29	−1.94	−38.67	−0.51	−1.30	−13.23	0.89	−25.44	−0.32	−0.95	−8812.78	1342.83	−1401.30	96.11	−1132.16	−9911.30	−2.29	−503.87

Khalifah RG. The carbon dioxide hydration activity of carbonic anhydrase. I. Stop-flow kinetic studies on the native human isoenzymes B and C. The Journal of Biological Chemistry 1971;246:2561–73

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