Journal: Biochemical and biophysical research communications

Article Title: Biophysical characterization of the structural change of Nopp140, an intrinsically disordered protein, in the interaction with CK2α

doi: 10.1016/j.bbrc.2016.06.040

Figure Lengend Snippet: (A) Schematic representation of Nopp140 and Nopp140Cs. Secondary structure regions of Nopp140 were predicted with the Quick2D bioinformatics toolkit (http://toolkit.tuebingen.mpg.de/quick2_d). Fluorescent dye labeled positions (S352C/S467C, S352/S589C, S352C/G660C, and S352C/S704C) are indicated with green asterisk symbol (*). EPR spins labeled positions (G568C, S589C, and S704C) are indicated with red arrows (↓). (B) SDS-PAGE analysis of MTSSL labeled Nopp140C (lane 1: before labeling, lane 2: after labeling) (C) SDS-PAGE analysis of Cy3-and Cy5-labeled Nopp140C. Images of lane 3 and lane 4 were obtained by fluorescence gel documentation. (lane 1: before labeling, lane 2: after labeling, lane 3: green laser excited Nopp140 C-terminus fragment, lane 4: red laser excited Nopp140 C-terminus fragment). (D) Representative absorbance spectrum of Cy3 and Cy5 labeled Nopp140C. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: The Nopp140C-S352C/S589C labeled with Cy3 and Cy5 in FRET buffer were immobilized onto a flow cell, in which a quartz slide was functionalized with PEG and PEG-biotin (Laysan Bio, 40:1).

Techniques: Labeling, SDS Page, Fluorescence

Journal: Biochemical and biophysical research communications

Article Title: Biophysical characterization of the structural change of Nopp140, an intrinsically disordered protein, in the interaction with CK2α

doi: 10.1016/j.bbrc.2016.06.040

Figure Lengend Snippet: Bulk fluorescence spectra of Cy3 and Cy5 labeled Nopp140Cs in the presence or absence of CK2α. With excitation at 550 nm, the spectra were obtained from 560 nm to 700 nm by with a fluorometer in the presence of 10 μM double-labeled Nopp140C (S352C/S467C) (A), Nopp140C(S352/S589C) (B), Nopp140C (S352C/G660C) (C), and Nopp140C (S352C/S704C) (D). Each spectrum of Nopp140C and Nopp140C with CK2α is presented with black and red dotted-lines, respectively. The molar ratio of Nopp140C-to-CK2α was 1:100. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: The Nopp140C-S352C/S589C labeled with Cy3 and Cy5 in FRET buffer were immobilized onto a flow cell, in which a quartz slide was functionalized with PEG and PEG-biotin (Laysan Bio, 40:1).

Techniques: Fluorescence, Labeling

Journal: Biochemical and biophysical research communications

Article Title: Biophysical characterization of the structural change of Nopp140, an intrinsically disordered protein, in the interaction with CK2α

doi: 10.1016/j.bbrc.2016.06.040

Figure Lengend Snippet: smFRET histograms and representative time series of double-labeled Nopp140C-S352C/S589C only (A, B), with CK2α (C, D), and in 6 M GdmCl (E, F). Histograms were fitted with up to two Gaussian peaks distributions having a FRET-peak centered at 0.03 (blue) and 0.45 E (red) each. Percentages of the areas of each Gaussian peak are indicated above each graph. The molar ratio of Nopp140C-to-CK2α was 1:100. For collecting the signals from Nopp140 labeled with both Cy3 and Cy5, an alternative excitation of Cy3 and Cy5 was performed using a 532 and 633 nm laser, respectively. Magenta arrows indicated intensity of Cy5 which was excited by 633 nm laser. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: The Nopp140C-S352C/S589C labeled with Cy3 and Cy5 in FRET buffer were immobilized onto a flow cell, in which a quartz slide was functionalized with PEG and PEG-biotin (Laysan Bio, 40:1).

Techniques: Labeling

Journal: Biochemical and biophysical research communications

Article Title: Biophysical characterization of the structural change of Nopp140, an intrinsically disordered protein, in the interaction with CK2α

doi: 10.1016/j.bbrc.2016.06.040

Figure Lengend Snippet: EPR spectra of spin-labeled Nopp140Cs (S568C, S589C, and S704C). The EPR spectra of spin labeled Nopp140Cs in the absence or presence of CK2α are presented as black and red dotted-lines, respectively. The spectra are normalized to the same integral for superimposition of data. The molar ratio of Nopp140C-to-CK2α was 1:100. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: The Nopp140C-S352C/S589C labeled with Cy3 and Cy5 in FRET buffer were immobilized onto a flow cell, in which a quartz slide was functionalized with PEG and PEG-biotin (Laysan Bio, 40:1).

Techniques: Labeling