Bioisis

"U2AF65 RRM1-RRM2"

Experimental SAS Curve

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Low_res_thumbnail

Experimental Mass

21,102 Da

Experimental Details for BID:  1U2FRP
Experiment ID: 90
Collected at: ALS BL 12.3.1
Contributors: Jenkins, JL ,  Kielkopf, CL ,  Laird, KM
SAXS profile of human U2AF65 RRM1-RRM2, residues 148-336, with N-terminal 'GPLGS' sequence from protease site. Sample was purified by gel filtration and homogeneity was verified by dynamic light scattering prior to data collection.
Samples at 2.6, 5, or 10 mg/mL were used for 6s and 60s exposures, followed by a 6s exposure to check for radiation damage. Appropriate resolution ranges of the exposures and concentrations were merged in the final data file.

Electron Pair Distribution

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       Dmax → 80 Å


Guinier Plot

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     Guinier Rg → 23.8 Å

Real Space Rg → 24.56 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.


Ensemble Model

A ENSEMBLE model was determined using the following:

Simulation MethodGAJOE 
Simulation AlgorithmGenetic Algorithm Judging Optimization of Ensembles 
Ensemble Size10000 
Selection MethodEOM 
Member Size
Scoring FunctionChi Square 
Score0.9 

Ensemble Fit

Med_res_ensemble_fit

The red line is the calculated SAXS profile from the ENSEMBLE model scaled to the experimental SAXS curve (cyan).


Additional Experimental Details
Title

U2AF65 RRM1-RRM2

Description

SAXS profile of human U2AF65 RRM1-RRM2, residues 148-336, with N-terminal 'GPLGS' sequence from protease site. Sample was purified by gel filtration and homogeneity was verified by dynamic light scattering prior to data collection.

Publication

(1) Jenkins JL, Shen H, Green MR, Kielkopf CL. J Biol Chem. (2008) Solution conformation and thermodynamic characteristics of RNA binding by the splicing factor U2AF65. 283(48):33641-9 (2) Major Contribution of Elongated Conformations to the Solution Ensemble of the Essential Splicing Factor U2AF65, submitted

Contributors

Jenkins, JL ,  Kielkopf, CL ,  Laird, KM

Genomics and Proteomics

The experiment is composed of a single gene/ORF

Abbreviated name: U2AF65R12

Annotation: RNA recognition motif domains RRM1 and RRM2 of essential pre-mRNA splicing factor U2AF65, with 'GPLGS' from cloning site

GPLGSARRLY VGNIPFGITE EAMMDFFNAQ MRLGGLTQAP GNPVLAVQIN QDKNFAFLEF RSVDETTQAM AFDGIIFQGQ SLKIRRPHDY QPLPGMSENP SVYVPGVVST VVPDSAHKLF IGGLPNYLND DQVKELLTSF GPLKAFNLVK DSATGLSKGY AFCEYVDINV TDQAIAGLNG MQLGDKKLLV QRAS
categoryamino acid composition(%)
HydrophobicI(4.6) V(7.7) L(10.3) M(3.1) A(7.7) G(9.8) P(6.2)
AromaticF(6.2) W(0.0) Y(3.1)
HydrophilicR(3.6) K(4.6) E(3.6) D(6.2) Q(6.2) N(5.7) H(1.0) S(5.7) T(4.1) C(0.5)