?url_ver=Z39.88-2004&rft_val_fmt=info%3Aofi%2Ffmt%3Akev%3Amtx%3Adc&rft.relation=http%3A%2F%2Fd-scholarship-dev.library.pitt.edu%2F10223%2F&rft.title=Insight+into+Structure+and+Function+of+EcoRI+Through+Detailed+Characterization+of+Nitroxide+Side+Chains+Using+Molecular+Dynamics+Simulations&rft.creator=Rajapakse%2C+Gayathri+Ayasna&rft.description=This+thesis+describes+molecular+dynamics+simulations+used+in+an+effort+to+investigate+the+orientation+and+dynamics+of+the+methane+thiosulfonate+spin+label+(MTSSL)+on+solvent+exposed+sites+of+the+loop+(R131C)+and+beta+strand+(S180C)+of+the+specific+EcoRI-DNA+complex.+Also%2C+the+results+are+compared+with+those+of+a+well-studied+solvent+exposed+alpha+helical+site+(SEHS).+By+modeling+the+spin+label+at+these+sites+using+the+all-atom+simulation%2C+we+intend+to+explain+the+observed+double+electron-electron+resonance+(DEER)+distance+distribution+data+of+the+specific+complex+of+R131C+and+S180C+mutants.+EcoRI+is+a+restriction+endonuclease%2C+which+binds+and+cleaves+the+specific+DNA+sequence+5'GAATTC3'.+The+arms+of+EcoRI+are+thought+to+play+an+important+role+in+its+high+binding+affinity.+However%2C+the+exact+reason(s)+why+it+shows+such+an+extraordinary+binding+specificity+is+still+being+investigated.+Therefore%2C+to+shed+light+on+the+high+binding+specificity+of+the+enzyme%2C+distance+measurements+were+previously+obtained+using+DEER%2C+providing+the+structural+constraints+of+the+arms+of+EcoRI.+Since+the+DEER+experiment+provides+inter-spin+label+distances%2C+simulations+can+be+used+to+de-convolute+the+spin+label+contribution+to+extract+the+inter-atomic+distances+(C-C)+of+the+arms+of+EcoRI.+In+order+to+accomplish+this%2C+we+intend+to+effectively+reproduce+the+experimental+DEER+data+using+molecular+dynamics+simulations.+Utilizing+simulations%2C+the+structural+constraints+contained+in+experimental+data+can+be+obtained+and+compared+with+the+X-ray+crystal+structure+of+the+specific+EcoRI-DNA+complex.+Although+we+attempt+to+explain+the+DEER+data+by+examining+spin-label+conformations%2C+the+study+also+involves+linking+the+dynamics+of+the+spin+label+to+the+MD-generated+distance+distributions.&rft.date=2011-01-27&rft.type=University+of+Pittsburgh+ETD&rft.type=PeerReviewed&rft.format=application%2Fpdf&rft.language=en&rft.identifier=http%3A%2F%2Fd-scholarship-dev.library.pitt.edu%2F10223%2F1%2FETD_Template_Rajapakse.pdf&rft.identifier=++Rajapakse%2C+Gayathri+Ayasna++(2011)+Insight+into+Structure+and+Function+of+EcoRI+Through+Detailed+Characterization+of+Nitroxide+Side+Chains+Using+Molecular+Dynamics+Simulations.++Master's+Thesis%2C+University+of+Pittsburgh.++++(Unpublished)++