Bioisis

"SAXS analysis of Hsp104 wild-type and N-terminal domain truncation"

Experimental SAS Curve

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Low_res_thumbnail

Experimental Mass

612,000 Da

Experimental Details for BID:  HS104P
Experiment ID: 112
Collected at: SSRL BL4-2, NSLS BL X-9
Contributors: Sweeny, EA ,  Jackrel, M ,  Sochor, MA ,  Razzo, B ,  Desantis, ME ,  Gupta, K ,  Shorter, J ,  Go , M
SAXS analysis of the yeast disaggregase Hsp104. Samples included the Wild-type (WT) and an N-terminal domain truncation (dN) each in 6 different nucleotide states; AMP-PNP, ATPgS, ATP, ADP-AlFx, ADP and no nucleotide. Prior to SAXS data collection each Hsp104 variant was purified with a final step of size-exclusion chromatography. Monodispersity was confirmed using multi-angle light scattering coupled to size exclusion chromatography.
Data was collected at concentrations between 1.5-6mg/mL.

Electron Pair Distribution

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       Dmax → 230 Å


Guinier Plot

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     Guinier Rg → 68.7 Å

Real Space Rg → 68.6 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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The Kratky plot can be 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.


GASBOR Model

GASBOR result was determined with the following:

Space GroupP6
Chi1.062
NSD | RMSD1.9 
variance(NSD | RMSD)0.13 
Number of Models in Average
Superimposed model

Additional Experimental Details
Title

SAXS analysis of Hsp104 wild-type and N-terminal domain truncation

Description

SAXS analysis of the yeast disaggregase Hsp104. Samples included the Wild-type (WT) and an N-terminal domain truncation (dN) each in 6 different nucleotide states; AMP-PNP, ATPgS, ATP, ADP-AlFx, ADP and no nucleotide. Prior to SAXS data collection each Hsp104 variant was purified with a final step of size-exclusion chromatography. Monodispersity was confirmed using multi-angle light scattering coupled to size exclusion chromatography.

Publication

The Hsp104 N-terminal Domain Enables Disaggregase Plasticity and Potentiation, Mol Cell.

Contributors

Sweeny, EA ,  Jackrel, M ,  Sochor, MA ,  Razzo, B ,  Desantis, ME ,  Gupta, K ,  Shorter, J ,  Go , M

Genomics and Proteomics

The experiment is composed of 2 genes/ORFs

Abbreviated name: Hsp104WT

Annotation: AAA+ disagregase protein from S. cerevisiae

MNDQTQFTER ALTILTLAQK LASDHQHPQL QPIHILAAFI ETPEDGSVPY LQNLIEKGRY DYDLFKKVVN RNLVRIPQQQ PAPAEITPSY ALGKVLQDAA KIQKQQKDSF IAQDHILFAL FNDSSIQQIF KEAQVDIEAI KQQALELRGN TRIDSRGADT NTPLEYLSKY AIDMTEQARQ GKLDPVIGRE EEIRSTIRVL ARRIKSNPCL IGEPGIGKTA IIEGVAQRII DDDVPTILQG AKLFSLDLAA LTAGAKYKGD FEERFKGVLK EIEESKTLIV LFIDEIHMLM GNGKDDAANI LKPALSRGQL KVIGATTNNE YRSIVEKDGA FERRFQKIEV AEPSVRQTVA ILRGLQPKYE IHHGVRILDS ALVTAAQLAK RYLPYRRLPD SALDLVDISC AGVAVARDSK PEELDSKERQ LQLIQVEIKA LERDEDADST TKDRLKLARQ KEASLQEELE PLRQRYNEEK HGHEELTQAK KKLDELENKA LDAERRYDTA TAADLRYFAI PDIKKQIEKL EDQVAEEERR AGANSMIQNV VDSDTISETA ARLTGIPVKK LSESENEKLI HMERDLSSEV VGQMDAIKAV SNAVRLSRSG LANPRQPASF LFLGLSGSGK TELAKKVAGF LFNDEDMMIR VDCSELSEKY AVSKLLGTTA GYVGYDEGGF LTNQLQYKPY SVLLFDEVEK AHPDVLTVML QMLDDGRITS GQGKTIDCSN CIVIMTSNLG AEFINSQQGS KIQESTKNLV MGAVRQHFRP EFLNRISSIV IFNKLSRKAI HKIVDIRLKE IEERFEQNDK HYKLNLTQEA KDFLAKYGYS DDMGARPLNR LIQNEILNKL ALRILKNEIK DKETVNVVLK KGKSRDENVP EEAEECLEVL PNHEATIGAD TLGDDDNEDS MEIDDDLD
categoryamino acid composition(%)
HydrophobicI(7.6) V(5.4) L(10.7) M(1.7) A(8.5) G(5.3) P(3.3)
AromaticF(2.9) W(0.0) Y(2.4)
HydrophilicR(5.9) K(7.7) E(9.0) D(7.5) Q(5.7) N(4.1) H(1.7) S(5.6) T(4.4) C(0.7)

Abbreviated name: Hsp104dN

Annotation: N-terminal domain truncation of the AAA+ protein disaggregase from S. cerevisiae Hsp104

EYLSKYAIDM TEQARQGKLD PVIGREEEIR STIRVLARRI KSNPCLIGEP GIGKTAIIEG VAQRIIDDDV PTILQGAKLF SLDLAALTAG AKYKGDFEER FKGVLKEIEE SKTLIVLFID EIHMLMGNGK DDAANILKPA LSRGQLKVIG ATTNNEYRSI VEKDGAFERR FQKIEVAEPS VRQTVAILRG LQPKYEIHHG VRILDSALVT AAQLAKRYLP YRRLPDSALD LVDISCAGVA VARDSKPEEL DSKERQLQLI QVEIKALERD EDADSTTKDR LKLARQKEAS LQEELEPLRQ RYNEEKHGHE ELTQAKKKLD ELENKALDAE RRYDTATAAD LRYFAIPDIK KQIEKLEDQV AEEERRAGAN SMIQNVVDSD TISETAARLT GIPVKKLSES ENEKLIHMER DLSSEVVGQM DAIKAVSNAV RLSRSGLANP RQPASFLFLG LSGSGKTELA KKVAGFLFND EDMMIRVDCS ELSEKYAVSK LLGTTAGYVG YDEGGFLTNQ LQYKPYSVLL FDEVEKAHPD VLTVMLQMLD DGRITSGQGK TIDCSNCIVI MTSNLGAEFI NSQQGSKIQE STKNLVMGAV RQHFRPEFLN RISSIVIFNK LSRKAIHKIV DIRLKEIEER FEQNDKHYKL NLTQEAKDFL AKYGYSDDMG ARPLNRLIQN EILNKLALRI LKNEIKDKET VNVVLKKGKS RDENVPEEAE ECLEVLPNHE ATIGADTLGD DDNEDSMEID DDLD
categoryamino acid composition(%)
HydrophobicI(7.3) V(5.8) L(10.8) M(1.9) A(8.2) G(5.8) P(2.8)
AromaticF(2.6) W(0.0) Y(2.4)
HydrophilicR(6.3) K(8.1) E(9.9) D(7.5) Q(4.3) N(4.0) H(1.5) S(5.9) T(4.2) C(0.8)