Inhibition Studies Of Antibiotics Resistant Target Protein Metallo-β-lactamases

In 1940,penicillin was put into use in clinical as the world’s first antibiotic.After 76 years of the development,antibiotics have saved millions of lives.However,with the wide use of antibiotics,antibiotic resistance has become an inevitable medical problems around the globe.β-Lactam antibiotics is one of the three largest clinical antibiotics at the moment.Many kinds of bacteria can produce β-lactamase which can hydrolyse β-lactam ring of these antibiotics,so that the P-lactam antibiotics lose their antibactericidal activity.β-Lactamase can be divided into four categories,among them,A,C and D class are serine(3-lactamase and they have clinical available inhibitors,but class B metallo-β-lactamase(MβLs)has no effective way to suppress due to its wide substrate range.This article mainly focuses on the inhibition of MβLs which is the antibiotic resistant target protein.Specific study contents are as follows:1.We express and purificate three subclasses of MβLs NDM-1,Ccr A,L1 and ImiS.It is a good foundation for inhibition kinetics research.2.We develop a series of azolylthioacetamide compounds as a new molecular skeleton structure of MβLs inhibitor.There are 42 azolylthioacetamide compounds which had been synthesized in two parts.First,we designed and obtained 18 compounds,and mainly discussed the effect of different azolyl and aromatic substituent on the inhibitory activity for the three subclasses MβLs.We find eventually that compounds contained thiazole ring showed better inhibitory activity against MβLs.Docking experiment showed how the compounds bind to the four kinds of MβLs.In order to study the structure-activity relationship,24 triazolylthioacetamide compounds were designed and synthesized.Through inhibition kinetics research,all the compounds showed a good inhibitory activity to NDM-1 with Ki value<2 μM.There is no obvious difference among different substituent in the two sides of triazole thioacetamide structure.Docking experiment confirms the important role of thiazolyl in inhibitory activity.3.β-Lactam ring is the basic structure of β-lactam antibiotics.Based on the character,a new cephalosporin compound ASC was designed and obtained.Through inhibition kinetics study,compound ASC can be seemed as a rare potential broad and competive MβLs inhibitor.It shows a better inhibitory activity for CcrA and ImiS than for NDM-1 and L1.ITC sduty shows the different rate that the four kinds of enzyme hydrolysize ASC,cefazolin sodium and imipenem and Km、kcat and ΔH are also obtained in the experiment.It shows that MβLs can hydrolysize ASC more quickly(2-6 times).In antibacterial activity experiment,compound ASC shows specific bactericidal effect on staphylococcus aureus’with MIC value of 256 μg/mL,and duration of the effect can be reached 40 hours or longer.In addition,compound ASC as inhibitor acted with clinical antibiotics can greatly reduce the usage of antibiotics and the largest reduction can reach 1000 times.4.Elements N and P are in the same group and phosphate/phosphate compounds also made some significant achievements in clinical drug application.Then based on the resistance mechanism of β-lactam antibiotics,we design a β-phosphoramide compounds so that we can radically solve the resistant status.We obtained 11 phosphate compounds which show inhibitory activity to MβLs Ccr A and NDM-1.We will try our best to optimize reaction conditions to obtain the(3-phosphoramide compound in the future.