Cellulase Linkers Are Optimized Based on Domain Type and Function Insights from Sequence Analysis, Biophysical Measurements, and Molecular Simulation 英文参考文献.docVIP

CellulaseLinkersAreOptimizedBasedonDomainType andFunction:InsightsfromSequenceAnalysis, BiophysicalMeasurements,andMolecularSimulation DeanneW.Sammond1,ChristinaM.Payne1,RomanBrunecky1,MichaelE.Himmel1,MichaelF.Crowley1, GreggT.Beckham2,3 * 1Biosciences Center, National Renewable Energy Laboratory, Golden, Colorado, United States of America, 2National Bioenergy Center, National Renewable Energy Laboratory,Golden,Colorado,UnitedStatesofAmerica,3DepartmentofChemicalEngineering,ColoradoSchoolofMines,Golden,Colorado,UnitedStatesofAmerica Abstract Cellulaseenzymesdeconstructcellulosetoglucose,andareoftencomprisedofglycosylatedlinkersconnectingglycoside hydrolases (GHs) to carbohydrate-binding modules (CBMs). Although linker modifications can alter cellulase activity, the functionalroleoflinkersbeyonddomainconnectivityremainsunknown.Hereweinvestigatecellulaselinkersconnecting GHFamily6or7catalyticdomainstoFamily1or2CBMs,frombothbacterialandeukaryoticcellulasestoidentifyconserved characteristics potentially related to function. Sequence analysis suggests that the linker lengths between structured domains are optimized based on the GH domain and CBM type, such that linker length may be important for activity. LongerlinkersareobservedineukaryoticGHFamily6cellulasescomparedtoGHFamily7cellulases.BacterialGHFamily6 cellulasesarefoundwithstructureddomainsineitherNtoCterminalorder,andsimilarlinkerlengthssuggestthereisno effect of domain order on length. O-glycosylation is uniformly distributed across linkers, suggesting that glycans are required along entire linker lengths for proteolysis protection and, as suggested by simulation, for extension. Sequence comparisonsshowthatprolinecontentforbacteriallinkersismorethandoublethatobservedineukaryoticlinkers,butwith fewerputativeO-glycansites,suggestingalternativemethodsforextension.Conversely,nearlinkerterminiwherelinkers connect to structured domains, O-glycosylation sites are observed less frequently, whereas glycines are more pr