ParameterizationofOPLS–AAForceFieldforthe ConformationalAnalysisofMacrocyclicPolyketides KALJUKAHN,THOMASC.BRUICE DepartmentofChemistryandBiochemistry,UniversityofCalifornia,SantaBarbara, SantaBarbara,California93106 Received27February2001;Accepted16November2001 Publishedonline00Month2002inWileyInterScience(www.interscience.wiley.com).DOI10.1002/jcc.10051 Abstract:TheparametersfortheOPLS–AApotentialenergyfunctionhavebeenextendedtoincludesomefunctional groupsthatarepresentinmacrocyclicpolyketides.ExistingOPLS–AAtorsionalparametersforalkanes,alcohols, ethers,hemiacetals,esters,andketoamideswereimprovedbasedonMP2/aug-cc-pVTZandMP2/aug-cc-pVDZ calculations.Nonbondedparametersforthesp3 carbonandoxygenatomswererefinedusingMonteCarlosimulations ofbulkliquids.Theresultingforcefieldpredictsconformerenergiesandtorsionalbarriersofalkanes,alcohols,ethers, andhemiacetalswithanoverallRMSdeviationof0.40kcal/molascomparedtoreferencedata.Densitiesof19bulk liquidsarepredictedwithanaverageerrorof1.1%,andheatsofvaporizationarereproducedwithin2.4%of experimentalvalues.Theforcefieldwasusedtoperformconformationalanalysisofsmalleranalogsofthemacrocyclic polyketidedrugFK506.Structuresthatadoptedlow-energyconformationssimilartothatofboundFK506were identified.Theresultsshowthatalinkeroffourketideunitsconstitutestheshortesteffectordomainthatallowsbinding oftheketidedrugstoFKBPproteins.Itisproposedthattheexactchemicalmakeupoftheeffectordomainhaslittle influenceontheconformationalpreferenceoftetraketides. ©2002WileyPeriodicals,Inc. JComputChem23:977–996,2002 Keywords:forcefielddevelopment;conformationalanalysis;neuroregenerativepolyketides Introduction ofcalcineurin,leadstosuppressionoftheimmunesystem.10 Thisis thebasisforcurrentimmunosuppressivetherapyusingFK506(also Polyketidesformastructurallydiverseclassofnaturalproducts knownasTacrolitmus).However,long-termsuppressionoftheim- whosebiosynthesistakesplacethroughthesuccessivecondensa- munesystemisundesirablewhiletreatingnerveinjury. 1–3 tionofsimplecarboxylicacids. Despitetheirwidelyvarying Weareinterestedinfindingpolyketidestructuresthatshow sizeandstructure,polyketidescanbegroupedintotwosuperfami- highaffinitytowardsFKBP52butdonotinteractwithcalcineurin. 2 lies:thearomaticandthecomplexpolyketides. Someexamplesof BindingstudieswithanalogsofFK506andavailablecrystal aromaticpolyketidesinclude6-methylsalicylicacidandtetracy- structuresofFKBP12⅐FK506⅐calcineurincomplexesrevealthat clines,whileerythromycinsbelongtothesuperfamilyofcomplex polyketidescanbeconsideredasbuiltfromthe“bindingdomain” polyketides.Manypolyketidesarepharmacologicallyactiveor thatinteractswithFKBP12,the“effectordomain”thatinteracts possessusefulagriculturalproperties.Itisthoughtthatthebiolog- withcalcineurin(shownboxedinChart1),andthecyclohexylarm icalactivityofcomplexpolyketidesisdependentonconforma- thatinteractsweaklywithbothproteins.10,11 Thestructureofthe tionalpreorganization,whichdirectsfunctionalgroupsinspace binaryFK506⅐FKBP12complexrevealsthatthepipecolinylester andreducestheentropicpenaltyinvolvedinbinding.4 moietyandthe ␣-ketoamide portion of the binding domain make Somecomplexmacrocyclicpolyketides,suchasFK506(Chart 1)anditsanalogs,arepromisingneuroregenerativeagents.5,6 Theirmechanismofactioncurrentlyispoorlyunderstood,but strongevidencesuggeststhatinteractionofapolyketidedrugwith Correspondenceto:T.C.Bruice;e-mail:[email protected] thecellularproteinFKBP52isrequiredforaccelerationofnerve Contract/grantsponsor:ChristopherReeveParalysisFoundation;contract/ cellgrowth.7 Asignificantprobleminusingpolyketidedrugsfor grantnumber:BA1-9904-2 treatmentofnerveinjuryisthatothercellularproteinsalsointeract Contract/grantsponsor:UniversityofMissouri-Columbia(K.K.) withthesemolecules.Theinteractionofapolyketidewithother Thisarticleincludessupplementarymaterialavailablefromtheauthorupon proteinscouldleadtounwantedsideeffects.Onewell-character- requestorviatheInternetatftp.wiley.com/public/journals/jcc/suppmat/ 8,9 izedtargetforFK506anditsanalogsistheproteinFKBP12. 23/977orhttp://www.interscience.wiley.com/jpages/0192-8651/suppmat/v23. TheinteractionofFK506withFKBP12,andthesubsequentbinding 977.html ©2002WileyPeriodicals,Inc. 978 Kahn and Bruice • Vol. 23, No. 10 • Journal of Computational Chemistry are similar and residues forming the binding pocket are highly conserved.18 The second approach consists of a design of mole- cules that would bind both FKBP52 and FKBP12 but have a modified effector domain rendering them unable to bind cal- cineurin. We assume, in the framework of this approach, that the binding modes of FK506 to the proteins FKBP12 and FKBP52 are similar. In this case, the problem reduces to finding polyketide structures that have low energy conformations with the binding domain maximally similar to the binding domain of the bound FK506. The viability of this approach has been demonstrated by evaluating the conformational energies of rapamycin analogs using the MM3 force field.19 In their work, Adalsteinsson and Bruice19 first performed a conformational search of three molecules (tem- plates) corresponding to the unsubstituted triketide, tetraketide, and pentaketide, respectively. Next, they calculated single point potential energies for a large number of substituted templates and Chart 1 identified structures that have low energy conformers similar to the conformation of the bound FK506. We now report an extension of this approach that allows a rapid conformational search with numerous favorable interactions with the FKBP12 protein.11,12 complete minimization of realistic polyketides as opposed to ide- The effector domain is not necessary for the polyketide to interact alized templates. with FKBP52, and various compounds lacking this linker region Computer modeling of biomolecular systems and large organic have been suggested as possible neuroregenerative drugs.13–15 One molecules is currently dominated by molecular mechanics calcu- shortcoming of drug candidates considered so far is that they can lations due to the advantage in speed. Several high quality force adopt a multitude of conformations, of which many are not com- fields, such as AMBER,20,21 CFF,22,23 CHARMM,24,25 COM- plementary with the binding site of FKBP52. In addition, many PASS,26 GROMOS,27 SPASIBA,28 MM3,29 MM4,30 OPLS– acyclic ligands bind to FKBP12 with a conformation that mini- AA,31 and MMFF9432,33 have been developed and are commonly mizes the amount of exposed hydrophobic surface area, a phenom- used for elucidating structures and properties of molecules in gas, enon called hydrophobic collapse.11 As a result, most acyclic liquid, and crystal phases. Currently, each force field has signifi- analogs show lower affinity towards FKBP12 than FK506 does.11 cant strength in the area where it was specifically parameterized, Also, the acyclic analogs of FK506 may require complicated that is, MMFF94 performs well for predicting the structures and organic synthesis. relative conformational energies of a wide range of organic mol- Studies with macrocyclic analogs of FK506 have shown that ecules while the OPLS–AA potential is well suited for describing many modifications in the effector domain abolish the interaction intermolecular interactions.32,34 Because we are also interested in with calcineurin while retaining a submicromolar affinity toward accurately predicting relative binding free energies of polyketides FKBP12. The comparison with FK506, which has a dissociation to target proteins via simulation methods, we chose the OPLS–AA constant of 0.4 nM from FKBP12,16 shows that further improve- force field for the conformational analysis. This force field has ment of the binding affinity should be possible. The macrocyclic been developed in the laboratory of William L. Jorgensen over the polyketides are especially promising candidates for neuroregen- course of several years and has been described in detail.31 In short, erative drugs because recent advances in identification and cloning the energy of a molecular system is derived as the sum of bond of polyketide synthase genes make it possible to produce such stretching, bond bending, torsional, and nonbonded terms. The polyketides via engineered biosynthesis.3 Such an approach prom- bond stretching and bending parameters come mostly from Wein- ises economic production of highly pure polyketides with the er’s 1986 AMBER force field,20 with the exception of alkane desired structure. In summary, the design goal for a neuroregen- parameters that have been adopted from CHARMM. The atomic erative drug is a compound that shows high affinity towards charges and Lennard–Jones parameters have been fitted to repro- FKBP52, does not interact with calcineurin, has a favorable solu- duce the densities, heats of vaporization, and free energies of bility profile, and can be synthesized economically via engineered hydration of a wide range of organic compounds.31,35–37 In some biosynthesis. instances, fitting to electrostatic potential has been used to assign In this article we have investigated which macrocyclic atomic charges.38 Also, ab initio calculations of intermolecular polyketide structures would bind strongly to FKBP52 but not energies of dimers or molecular hydrates in the gas phase have interact with calcineurin. In principle, at least two approaches are been used for determination of nonbonded parameters.35,38 Most possible for designing such compounds. In the first approach, one torsional parameters in the OPLS–AA force field were derived could try to design molecules that bind strongly to FKBP52 and do based on HF/6-31G* calculations