Study On Crystal Structure And Enzyme Activity Of Inducible Lactate Dehydrogenase From Staphylococcus Aureus

Staphylococcus aureus (S.aureus)is one of the most prevalent pathogens tocause human infection.Pathogenic S.aureus is resistant to human innate immuneantimicrobe NO*,which is the first line of host defense depending on the mechanismthat NO* can inhibit bacterial respiration.Study indicated that S.aureus can changemetabolic state to survive from nitrosative stress,when a NO* inducible l-lactatedehydrogenase (ldh-1 SACOL0222)is expressed,which is also essential for thebacterial virulence.Thus,the inducible 1-lactate dehydrogenase in S.aureus wouldbecome a potential target for new class of antibiotics if we can find a selectiveinhibitor,which can potently inhibit S.aureus ldhl but shows much no inhibitionactivity against the human LDH isoforms.To insight into its innate immune mechanism and to screen selective inhibitorbased structure for potential drug discovery,resoving the 3 dimentional structure ofldh1 from S.aureus is the key step.This thesis describes the l-lactate dehydrogenasestructure with 2.2 A resolution and the enzymatic kenetics study of converting lactateto pyruvate with NAD coenzyme.The crystals diffracted belonged to space group C2,and the unit-cell parameters are a=131.36,b=74.362,c=103.243.The final model ofldh-1 from S.aureus is refined to 2.18 A with R-factor and freer-factor of 17% and23.4% respectively.There are two molecules per asymmetric unit (shown in Figure 3).There are two domains:the larger with a"Rossmann"type fold formed by residue22-160 and 248-264.Active site loop of Idh-1 from S.aureus is highly conserved(96-108)between spcies,however,adopts a more open conformation,which increasethe volume of the active site cleft in ldh-1 from S.aureus.From enzymetic keneticsstudy,significant increase of Km and Kcat feetures ldh-1 from human isoforms.