"Solution structure of DNA-PKcs"

Experimental Scattering Curve
Med_res_iofq

Low_res_thumbnail

Experimental Mass

497,000 Da
Experimental Details for BID:  1DPKCY
Experiment ID: 51
Collected at: ALS BL 12.3.1
Contributors: Hammel, M. ,  Mahaney, B.L. ,  Leese-Miller, S.P. ,  Tainer, J.A.
The SAXS curves displayed a concentration-dependence arising from the tendency of DNA-PKcs to undergo self-association at high protein concentrations. Interference-free SAXS profiles were estimated by extrapolating the measured scattering curves to infinite dilution. Analysis of the interference free SAXS profile, which represents the monomeric state of DNA-PKcs, shows a typical globular particle with a Dmax of ~155 Angstroms. Although variation exists between the individual SAXS envelopes, the position of the head and palm regions previously described in cryo-EM envelopes and crystal structure could be visualized. Similar to the 3D-EM reconstructions, the SAXS envelopes show distinct hollow features located inside the head region. Our SAXS envelopes show flattening of the shape in one direction which is consistent with the structure determined crystallography. To test the accuracy of the DNA-PKcs shape, we measured a second independently prepared sample. The P(r) functions matched well and the reconstructed average shapes show identical structural features in handedness symmetry in both possible hands as expected.
Merged from several concentrations between 1 and 15 mg/mL

Electron Pair Distribution

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Med_res_pofr

       Dmax → 155 Å

Guinier Plot

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Med_res_guinier

     Guinier Rg → 55.0 Å

Real Space Rg → 54.45 Å

The Guinier plot is used to estimate the radius of gyration, Rg, which is taken from the slope of a line observed at low scattering angles (typically in the range where q* Rg < 1.3). This should be in reasonable agreement with the real space Rg.

Kratky Plot

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Med_res_kratky

The Kratky plot is used to visually assess the degree of "unfoldedness" of a protein or RNA sample. The plot of a well-behaved folded protein approaches the baseline at high q values creating a parabolic shape similar to that for the Electron Pair Distribution Plot.

GASBOR Model

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GASBOR result was determined with the following:

Space GroupP1
Chi1.0
NSD | RMSD1.79 
variance(NSD | RMSD)0.01 
Number of Models in Average10 
Superimposed model10 
Additional Experimental Details
Title

Solution structure of DNA-PKcs

Description

The SAXS curves displayed a concentration-dependence arising from the tendency of DNA-PKcs to undergo self-association at high protein concentrations. Interference-free SAXS profiles were estimated by extrapolating the measured scattering curves to infinite dilution. Analysis of the interference free SAXS profile, which represents the monomeric state of DNA-PKcs, shows a typical globular particle with a Dmax of ~155 Angstroms. Although variation exists between the individual SAXS envelopes, the position of the head and palm regions previously described in cryo-EM envelopes and crystal structure could be visualized. Similar to the 3D-EM reconstructions, the SAXS envelopes show distinct hollow features located inside the head region. Our SAXS envelopes show flattening of the shape in one direction which is consistent with the structure determined crystallography. To test the accuracy of the DNA-PKcs shape, we measured a second independently prepared sample. The P(r) functions matched well and the reconstructed average shapes show identical structural features in handedness symmetry in both possible hands as expected.

Publication

DNA-dependent protein kinase dynamic conformations and assembly regulate DNA binding and the initial non-homologous end joining complex. J Biol Chem. 2010;285(2):1414-23. PMCID: 2801267.

Contributors

Hammel, M. ,  Mahaney, B.L. ,  Leese-Miller, S.P. ,  Tainer, J.A.

Genomics and Proteomics

The experiment is composed of 2 genes/ORFs

Abbreviated name: HUMDNAPKCS

Annotation: HUMAN DNA-dependent protein kinase catalytic subunit

MAGSGAGVRC SLLRLQETLS AADRCGAALA GHQLIRGLGQ ECVLSSSPAV LALQTSLVFS RDFGLLVFVR KSLNSIEFRE CREEILKFLC IFLEKMGQKI APYSVEIKNT CTSVYTKDRA AKCKIPALDL LIKLLQTFRS SRLMDEFKIG ELFSKFYGEL ALKKKIPDTV LEKVYELLGL LGEVHPSEMI NNAENLFRAF LGELKTQMTS AVREPKLPVL AGCLKGLSSL LCNFTKSMEE DPQTSREIFN FVLKAIRPQI DLKRYAVPSA GLRLFALHAS QFSTCLLDNY VSLFEVLLKW CAHTNVELKK AALSALESFL KQVSNMVAKN AEMHKNKLQY FMEQFYGIIR NVDSNNKELS IAIRGYGLFA GPCKVINAKD VDFMYVELIQ RCKQMFLTQT DTGDDRVYQM PSFLQSVASV LLYLDTVPEV YTPVLEHLVV MQIDSFPQYS PKMQLVCCRA IVKVFLALAA KGPVLRNCIS TVVHQGLIRI CSKPVVLPKG PESESEDHRA SGEVRTGKWK VPTYKDYVDL FRHLLSSDQM MDSILADEAF FSVNSSSESL NHLLYDEFVK SVLKIVEKLD LTLEIQTVGE QENGDEAPGV WMIPTSDPAA NLHPAKPKDF SAFINLVEFC REILPEKQAE FFEPWVYSFS YELILQSTRL PLISGFYKLL SITVRNAKKI KYFEGVSPKS LKHSPEDPEK YSCFALFVKF GKEVAVKMKQ YKDELLASCL TFLLSLPHNI IELDVRAYVP ALQMAFKLGL SYTPLAEVGL NALEEWSIYI DRHVMQPYYK DILPCLDGYL KTSALSDETK NNWEVSALSR AAQKGFNKVV LKHLKKTKNL SSNEAISLEE IRIRVVQMLG SLGGQINKNL LTVTSSDEMM KSYVAWDREK RLSFAVPFRE MKPVIFLDVF LPRVTELALT ASDRQTKVAA CELLHSMVMF MLGKATQMPE GGQGAPPMYQ LYKRTFPVLL RLACDVDQVT RQLYEPLVMQ LIHWFTNNKK FESQDTVALL EAILDGIVDP VDSTLRDFCG RCIREFLKWS IKQITPQQQE KSPVNTKSLF KRLYSLALHP NAFKRLGASL AFNNIYREFR EEESLVEQFV FEALVIYMES LALAHADEKS LGTIQQCCDA IDHLCRIIEK KHVSLNKAKK RRLPRGFPPS ASLCLLDLVK WLLAHCGRPQ TECRHKSIEL FYKFVPLLPG NRSPNLWLKD VLKEEGVSFL INTFEGGGCG QPSGILAQPT LLYLRGPFSL QATLCWLDLL LAALECYNTF IGERTVGALQ VLGTEAQSSL LKAVAFFLES IAMHDIIAAE KCFGTGAAGN RTSPQEGERY NYSKCTVVVR IMEFTTTLLN TSPEGWKLLK KDLCNTHLMR VLVQTLCEPA SIGFNIGDVQ VMAHLPDVCV NLMKALKMSP YKDILETHLR EKITAQSIEE LCAVNLYGPD AQVDRSRLAA VVSACKQLHR AGLLHNILPS QSTDLHHSVG TELLSLVYKG IAPGDERQCL PSLDLSCKQL ASGLLELAFA FGGLCERLVS LLLNPAVLST ASLGSSQGSV IHFSHGEYFY SLFSETINTE LLKNLDLAVL ELMQSSVDNT KMVSAVLNGM LDQSFRERAN QKHQGLKLAT TILQHWKKCD SWWAKDSPLE TKMAVLALLA KILQIDSSVS FNTSHGSFPE VFTTYISLLA DTKLDLHLKG QAVTLLPFFT SLTGGSLEEL RRVLEQLIVA HFPMQSREFP PGTPRFNNYV DCMKKFLDAL ELSQSPMLLE LMTEVLCREQ QHVMEELFQS SFRRIARRGS CVTQVGLLES VYEMFRKDDP RLSFTRQSFV DRSLLTLLWH CSLDALREFF STIVVDAIDV LKSRFTKLNE STFDTQITKK MGYYKILDVM YSRLPKDDVH AKESKINQVF HGSCITEGNE LTKTLIKLCY DAFTENMAGE NQLLERRRLY HCAAYNCAIS VICCVFNELK FYQGFLFSEK PEKNLLIFEN LIDLKRRYNF PVEVEVPMER KKKYIEIRKE AREAANGDSD GPSYMSSLSY LADSTLSEEM SQFDFSTGVQ SYSYSSQDPR PATGRFRRRE QRDPTVHDDV LELEMDELNR HECMAPLTAL VKHMHRSLGP PQGEEDSVPR DLPSWMKFLH GKLGNPIVPL NIRLFLAKLV INTEEVFRPY AKHWLSPLLQ LAASENNGGE GIHYMVVEIV ATILSWTGLA TPTGVPKDEV LANRLLNFLM KHVFHPKRAV FRHNLEIIKT LVECWKDCLS IPYRLIFEKF SGKDPNSKDN SVGIQLLGIV MANDLPPYDP QCGIQSSEYF QALVNNMSFV RYKEVYAAAA EVLGLILRYV MERKNILEES LCELVAKQLK QHQNTMEDKF IVCLNKVTKS FPPLADRFMN AVFFLLPKFH GVLKTLCLEV VLCRVEGMTE LYFQLKSKDF VQVMRHRDDE RQKVCLDIIY KMMPKLKPVE LRELLNPVVE FVSHPSTTCR EQMYNILMWI HDNYRDPESE TDNDSQEIFK LAKDVLIQGL IDENPGLQLI IRNFWSHETR LPSNTLDRLL ALNSLYSPKI EVHFLSLATN FLLEMTSMSP DYPNPMFEHP LSECEFQEYT IDSDWRFRST VLTPMFVETQ ASQGTLQTRT QEGSLSARWP VAGQIRATQQ QHDFTLTQTA DGRSSFDWLT GSSTDPLVDH TSPSSDSLLF AHKRSERLQR APLKSVGPDF GKKRLGLPGD EVDNKVKGAA GRTDLLRLRR RFMRDQEKLS LMYARKGVAE QKREKEIKSE LKMKQDAQVV LYRSYRHGDL PDIQIKHSSL ITPLQAVAQR DPIIAKQLFS SLFSGILKEM DKFKTLSEKN NITQKLLQDF NRFLNTTFSF FPPFVSCIQD ISCQHAALLS LDPAAVSAGC LASLQQPVGI RLLEEALLRL LPAELPAKRV RGKARLPPDV LRWVELAKLY RSIGEYDVLR GIFTSEIGTK QITQSALLAE ARSDYSEAAK QYDEALNKQD WVDGEPTEAE KDFWELASLD CYNHLAEWKS LEYCSTASID SENPPDLNKI WSEPFYQETY LPYMIRSKLK LLLQGEADQS LLTFIDKAMH GELQKAILEL HYSQELSLLY LLQDDVDRAK YYIQNGIQSF MQNYSSIDVL LHQSRLTKLQ SVQALTEIQE FISFISKQGN LSSQVPLKRL LNTWTNRYPD AKMDPMNIWD DIITNRCFFL SKIEEKLTPL PEDNSMNVDQ DGDPSDRMEV QEQEEDISSL IRSCKFSMKM KMIDSARKQN NFSLAMKLLK ELHKESKTRD DWLVSWVQSY CRLSHCRSRS QGCSEQVLTV LKTVSLLDEN NVSSYLSKNI LAFRDQNILL GTTYRIIANA LSSEPACLAE IEEDKARRIL ELSGSSSEDS EKVIAGLYQR AFQHLSEAVQ AAEEEAQPPS WSCGPAAGVI DAYMTLADFC DQQLRKEEEN ASVIDSAELQ AYPALVVEKM LKALKLNSNE ARLKFPRLLQ IIERYPEETL SLMTKEISSV PCWQFISWIS HMVALLDKDQ AVAVQHSVEE ITDNYPQAIV YPFIISSESY SFKDTSTGHK NKEFVARIKS KLDQGGVIQD FINALDQLSN PELLFKDWSN DVRAELAKTP VNKKNIEKMY ERMYAALGDP KAPGLGAFRR KFIQTFGKEF DKHFGKGGSK LLRMKLSDFN DITNMLLLKM NKDSKPPGNL KECSPWMSDF KVEFLRNELE IPGQYDGRGK PLPEYHVRIA GFDERVTVMA SLRRPKRIII RGHDEREHPF LVKGGEDLRQ DQRVEQLFQV MNGILAQDSA CSQRALQLRT YSVVPMTSRL GLIEWLENTV TLKDLLLNTM SQEEKAAYLS DPRAPPCEYK DWLTKMSGKH DVGAYMLMYK GANRTETVTS FRKRESKVPA DLLKRAFVRM STSPEAFLAL RSHFASSHAL ICISHWILGI GDRHLNNFMV AMETGGVIGI DFGHAFGSAT QFLPVPELMP FRLTRQFINL MLPMKETGLM YSIMVHALRA FRSDPGLLTN TMDVFVKEPS FDWKNFEQKM LKKGGSWIQE INVAEKNWYP RQKICYAKRK LAGANPAVIT CDELLLGHEK APAFRDYVAV ARGSKDHNIR AQEPESGLSE ETQVKCLMDQ ATDPNILGRT WEGWEPWM
categoryamino acid composition(%)
HydrophobicI(4.8) V(6.1) L(12.8) M(2.9) A(6.5) G(4.5) P(4.5)
AromaticF(4.7) W(1.2) Y(2.9)
HydrophilicR(5.2) K(6.7) E(7.2) D(5.1) Q(4.6) N(3.7) H(2.2) S(7.9) T(4.4) C(2.1)

Abbreviated name: DNAHAIRPIN

Annotation: 40 bp stem loop DNA

GCTACTTGTT GCTATCATCT GCTTTCGTTG TTACCTCATT AGCTTTCGAT GAACAACGAT AGTAGACGAA AGCAACAATG GAGTAATCGA CC
categoryResidue composition (%)
Nucleotides G (19.6) C (21.7) A (28.3) U (0.0) T (30.4)
Mass: 28,449 Da | Total residues: 92