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Low Activity Oligomers of Porphobilinogen Synthase as Antibiotic Targets

作为抗生素靶标的胆色素原合酶的低活性寡聚物

基本信息

批准号：
8069778
负责人：
EILEEN K JAFFE
金额：
$ 1.07万
依托单位：
RESEARCH INST OF FOX CHASE CAN CTR
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-09-23 至 2010-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8069778
关键词：
Active Sites Allosteric Regulation Anabolism Antibiotics Binding Binding Sites Biological Assay Biological Models Cancer Patient Cell Culture System Cell Culture Techniques Cessation of life Childhood Collaborations Complex Computer Simulation Core Facility DNA Sequence Rearrangement Developing Countries Development Disease Docking Encephalitis Enzymes Equilibrium Gelshift Analysis Heme Homo Hospitals Human Immune In Vitro Individual Infection Kinetics Knowledge Laboratories Libraries Malaria Methods Microbe Modeling Morphologic artifacts National Institute of Allergy and Infectious Disease Organism Parasites Phylogenetic Analysis Plants Plasmodium falciparum Porphobilinogen Synthase Positioning Attribute Postdoctoral Fellow Preparation Process Protein Isoforms Proteins Pseudomonas aeruginosa Relative (related person)Resources Roentgen Rays Scheme Structure Surface Technology Testing Tetrapyrroles Therapeutic Agents Toxoplasma gondii Variant Vibrio cholerae Virulence Work X-Ray Crystallography abstracting analog antimicrobial drug base cofactor comparative cystic fibrosis patients drug development drug discovery enzyme activity expression cloning in vitro testing in vivo inhibitor/antagonist non-drug pathogen protein oligomer protein structure small molecule virtual

项目摘要

Abstract: Allosteric regulation through the interconversion of functionally distinct non-additive quaternary structure assemblies has recently been described for the essential enzyme porphobilinogen synthase (PBGS). The oligomeric rearrangements seen with PBGS are different from the MWC or Koshland models for allostery; thus, the term morpheein has been introduced to describe this structural basis for allostery. Small molecules that can stabilize either an active or inactive quaternary structure assembly, herein called morphlocks, are proposed to function as therapeutic agents. Stabilization of an inactive quaternary structure assembly of porphobilinogen synthase (PBGS) is proposed as the basis for a new class of antibiotic agents. Although PBGS, which catalyzes the first common step in the biosynthesis of the tetrapyrrole cofactors (e.g. heme), is essential in both humans and most human pathogens, there is extensive phylogenetic variation in the putative morphlock binding sites. This allows us to target PBGS for drug development. There are four Specific Aims. Aim 1 is the application of in silico methods (protein structure modeling and docking) to the discovery of morphlocks that will trap inactive quaternary structure assemblies of PBGS from the human pathogens Pseudomonas aeruginosa, Vibrio cholera, Toxoplasma gondii, and Plasmodium falciparum. Post-docking criteria will be used to select compounds for purchase. Aim 2 is the in vitro testing of the purchased compounds as inhibitors of PBGS from the targeted pathogens. Using native gel shift analysis and extensive kinetic criteria, purchased compounds will be carefully and critically evaluated to select inhibitors that function through the oligomer stabilization mechanism and with species selectivity. Aim 3 is the in vivo testing of chosen morphlocks using a human cell culture system and standard microbiological assays. In Aim 4 we will use X-ray crystallography to evaluate the structures of the morphlock-PBGS complexes. To further probe the structure/function of select morphlocks, we will synthesize and evaluate structural analogs. The proposed methods have been used to discover the first morphlock, morphlock-1, which inhibits a plant PBGS in a species selective fashion by trapping the inactive hexameric assembly. Should the proposed morpheein mechanism for allosteric regulation be applicable to other medically important proteins, the proposed work will serve as a roadmap for harnessing this allosteric mechanism for drug discovery.

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