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An efficient strategy to enhance binding affinity and specificity of a known isozyme inhibitor.

ABSTRACT: The binding profile of a known inhibitor, benzenesulfonamide, against a family of carbonic anhydrase isozymes was efficiently enhanced via high-throughput screening of customized combinatorial one-bead-one-compound peptide libraries modified with the inhibitor molecule. The screening of the conjugate libraries recognized subtle variations in the microenvironments of the target enzyme and thus facilitated the identification of short peptide sequences that bind selectively to a close proximity of the active site. The identified peptide portions contributed significantly to the overall binding of the conjugate peptides with greatly enhanced affinity as well as improved specificity towards the target isozyme. The interactions between the inhibitors and the isozymes were validated by surface plasmon resonance (SPR), pull-down assay and enzymatic activity measurement. This high-throughput approach proved useful and efficient to enhance the binding profile of known inhibitors and may apply to developing effective inhibitors for a wide range of isozyme families.

SUBMITTER: Jee JE

PROVIDER: S-EPMC4942345 | biostudies-literature | 2016 Jul

REPOSITORIES: biostudies-literature

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An efficient strategy to enhance binding affinity and specificity of a known isozyme inhibitor.

Jee Joo-Eun JE Lim Jaehong J Ong Yong Siang YS Oon Jessica J Gao Liqian L Choi Hak Soo HS Lee Su Seong SS

Organic & biomolecular chemistry 20160701 28

The binding profile of a known inhibitor, benzenesulfonamide, against a family of carbonic anhydrase isozymes was efficiently enhanced via high-throughput screening of customized combinatorial one-bead-one-compound peptide libraries modified with the inhibitor molecule. The screening of the conjugate libraries recognized subtle variations in the microenvironments of the target enzyme and thus facilitated the identification of short peptide sequences that bind selectively to a close proximity of ...[more]

PMID: 27339902

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Project description:Dihydroneopterin aldolase (DHNA) is essential for folate biosynthesis in microorganisms. Without a counterpart in mammals, DHNA is an attractive target for antimicrobial agents. Helicobacter pylori infection occurs in human stomach of over 50% of the world population, but first-line therapies for the infection are facing rapidly increasing resistance. Novel antibiotics are urgently needed, toward which structural information on potential targets is critical. We have determined the crystal structure of H. pylori DHNA (HpDHNA) in complex with a pterin molecule (HpDHNA:Pterin) at 1.49-Å resolution. The HpDHNA:Pterin complex forms a tetramer in crystal. The tetramer is also observed in solution by dynamic light scattering and confirmed by small-angle X-ray scattering. To date, all but one reported DHNA structures are octameric complexes. As the only exception, ligand-free Mycobacterium tuberculosis DHNA (apo-MtDHNA) forms a tetramer in crystal, but its active sites are only partially formed. In contrast, the tetrameric HpDHNA:Pterin complex has well-formed active sites. Each active site accommodates one pterin molecule, but the exit of active site is blocked by two amino acid residues exhibiting a contact distance of 5.2 Å. In contrast, the corresponding contact distance in Staphylococcus aureus DHNA (SaDHNA) is twice the size, ranging from 9.8 to 10.5 Å, for ligand-free enzyme, the substrate complex, the product complex, and an inhibitor complex. This large contact distance indicates that the active site of SaDHNA is wide open. We propose that this isozyme-specific contact distance (ISCD) is a characteristic feature of DHNA active site. Comparative analysis of HpDHNA and SaDHNA structures suggests a fragment-based strategy for the development of isozyme-specific inhibitors. Graphical abstract Image 1 Highlights • Crystal structure of Helicobacter pylori DHNA in complex with pterin at 1.49 Å.• H. pylori DHNA exists as a tetramer in crystal with well-formed active sites.• The tetrameric H. pylori DHNA assembly also exits in solution.• Structural comparison reveals an isozyme-specific feature of DHNA active sites.• H. pylori DHNA structure suggests a strategy for isozyme-specific inhibitor design.

Dataset Information

An efficient strategy to enhance binding affinity and specificity of a known isozyme inhibitor.

Publications

An efficient strategy to enhance binding affinity and specificity of a known isozyme inhibitor.

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