Crystal structure of the NanR and ManNAc-6P complex, and uses thereof

Patent metadata

Field	Value
Publication number	US-9482669-B2
Application number	US-201414310879-A
Country	US
Kind code	B2
Filing date	Jun 20, 2014
Priority date	Jun 21, 2013
Publication date	Nov 1, 2016
Grant date	Nov 1, 2016

Abstract

The present invention relates to a three-dimensional structure of a complex explored by crystallization of the complex of NanR which is a key pathogenic regulatory protein of Vibrio vulnificus and ManNA6P which is a NanR regulator. Further, the present invention relates to a modified NanR protein, a polynucleotide encoding the protein, a vector including the polynucleotide, and a transformant including the vector. Furthermore, the present invention relates to a method for screening a substance regulating interaction between NanR and the transcriptional control region of nan operon which is a gene cluster regulated by NanR, or a substance regulating interaction between NanR and ManNAc-6P, by designing three-dimensional structure of the complex, and to an antibacterial composition including the screened substance.

First claim

What is claimed is:

1. A method for screening a substance regulating the interaction between NanR and the transcriptional control region of a nan operon, comprising
(a) designing and displaying a tertiary structure of a complex of NanR protein and ManNAc-6P using the atomic coordinates of the complex shown in Table 3,
wherein the complex comprises binding site amino acid residues R71, A137, S138, H163, S182, S183, S184, T187, E229, P231, G234 and K240;
(b) preparing candidate substances binding to NanR by employing the identified binding site amino acids from (a) to design and fit said substances; and
(c) examining binding affinity of the candidate substances for NanR and its regulation of the interaction between NanR and the transcriptional control region of nan operon,
wherein step (a) is carried out using a computer, and
wherein the atomic coordinates of the complex shown in Table 3 are stored on non-transitory electronic media that is used by the computer.

2. The method according to claim 1 , further comprising determining the candidate as a bacterial growth inhibitor, when the candidate binding to NanR has the NanR binding affinity similar to or higher than that of ManNAc-6P and maintains or increases interaction between NanR and the transcriptional control region of nan operon, compared to a control group treated without the corresponding candidate.

3. The method according to claim 1 , further comprising determining the candidate as a bacterial growth stimulant, when the candidate binding to NanR has the NanR binding affinity similar to or higher than that of ManNAc-6P and decreases interaction between NanR and the transcriptional control region of nan operon, compared to a control group treated without the corresponding candidate.

4. A method for screening a substance regulating the interaction between NanR and ManNAc-6P, comprising
(a) designing and displaying a tertiary structure of a complex of NanR protein and ManNAc-6P using the atomic coordinates of the complex shown in Table 3,
wherein the complex comprises binding site amino acid residues R71, A137, S138, H163, S182, S183, S184, T187, E229, P231, G234 and K240;
(b) preparing candidate substances binding to NanR by employing the identified binding site amino acids from (a) to design and fit said substances; and
(c) examining whether the candidate regulates the interaction between NanR protein and ManNAc-6P,
wherein step (a) is carried out using a computer, and
wherein the atomic coordinates of the complex shown in Table 3 are stored on non-transitory electronic media that is used by the computer.

5. The method according to claim 4 , further comprising determining the candidate as a nan operon expression enhancer or as a bacterial growth stimulant, when the candidate increases interaction between NanR and ManNAc-6P, compared to a control group treated without the corresponding candidate.

6. The method according to claim 4 , further comprising determining the candidate as a nan operon expression suppressor or as a bacterial growth inhibitor, when the candidate decreases interaction between NanR and ManNAc-6P, compared to a control group treated without the corresponding candidate.

Assignees

• Snu R&Db Foundation
• Korea Res Inst Bioscience & Biotechnology

Inventors

• Choi Sang Ho
• Kim Byoung Sik
• Kim Myung Hee
• Hwang Jung Won

Classifications

• G01N2500/04

  Screening involving studying the effect of compounds C directly on molecule A (e.g. C are potential ligands for a receptor A, or potential substrates for an enzyme A) · CPC title

• G01N33/56911 Primary

  Bacteria · CPC title

• C07K14/28

  from Vibrionaceae (F) · CPC title

• G01N2400/00

  Assays, e.g. immunoassays or enzyme assays, involving carbohydrates · CPC title