1234567891011 His-eIF5A Lys Elution 50 20 FTL Online Resource Figure 1. Purification of 6xHis-eIF5A Lys from E. coli. E. coli strain M15 was transformed.

Slides:

Advertisements

Similar presentations

Supplementary figure 1:

Advertisements

Figure Essential Cell Biology (© Garland Science 2010)

Helicobacter pylori Expresses an Autolytic Enzyme: Gene Identification, Cloning, and Theoretical Protein Structure Eleonora Marsich,1 Pierfrancesco Zuccato,1.

Ameer Effat M. Elfarash Dept. of Genetics Fac. of Agriculture, Assiut Univ. Gene Expression.

Expression Vector Expression of cloned genes produces large quantities of protein Components of expression vector 1. replication origin 2. polylingker.

pGLO™ Transformation and Purification of

Protein Purification and Expression MCB 130L, Lecture 2.

Protein Purification and Expression

Protein Purification. What do you know about proteins? Why do we need to purify proteins? What are you curious about?

4 September, 2006 Chapters Methods: Proteins, Model Systems I.

Protein Purification. Why purify Proteins? Characterize Function Activity Structure Study protein regulation and protein interactions Use in assays Produce.

Molecular Cell Biology Purifying Proteins Cooper.

SUPPLEMENTARY MATERIAL consist of two figures: Figure S1 contains two SDS-PAGE gels showing the purification profile of the recombinant yeast and slime.

MACS® Technology for magnetic isolation of cells and molecules

Plasmids Indispensable tools that allow molecular biologists to obtain essentially unlimited amounts of a DNA sequence Small circular DNA molecules that.

Figure 5: Expression and solubility tests for constructs of CoVs. Coronaviruses are complex, positive-sense RNA viruses that cause mild to severe respiratory.

pGLO™ Transformation and Purification of Green Fluorescent Protein (GFP)

Protein Overexpression in E. coli and

Protein Overexpression in E

Protein Overexpression in E. coli and

Figure Molecular Biology of the Cell (© Garland Science 2008)

Figure 14-1 Molecular Biology of the Cell (© Garland Science 2008)

Figure S1. Production of recombinant NS1 protein

Evaluation of a Highly Skin Permeable Low-Molecular-Weight Protamine Conjugated Epidermal Growth Factor for Novel Burn Wound Healing Therapy Ji Hae Lee,

Fac. of Agriculture, Assiut Univ.

Target protein Additional file 3. SDS-PAGE showing the degree of purification of D1-26PtxtPL1-27 expressed in E. coli. PtxtPL1-27.

VWF73, a region from D1596 to R1668 of von Willebrand factor, provides a minimal substrate for ADAMTS-13 by Koichi Kokame, Masanori Matsumoto, Yoshihiro.

Affinity Chromatography

Purification of recombinant T7 RNA polymerase

BRCA1 Is Associated with a Human SWI/SNF-Related Complex

Timur M. Yusufzai, Hideaki Tagami, Yoshihiro Nakatani, Gary Felsenfeld

Human peptidoglycan recognition protein S is an effector of neutrophil-mediated innate immunity by Ju Hyun Cho, Iain P. Fraser, Koichi Fukase, Shoichi.

Volume 14, Issue 9, Pages (September 2007)

Hours after rec PR-Set7 release Ladder kDa 15 PR-Set7 Coomassie

H-NS2 protein interacts with Lon and H-NS protein in vitro.

Glen S. Cho, Jack W. Szostak Chemistry & Biology

Protein purification and quality control.

Matthew D. Petroski, Raymond J. Deshaies Molecular Cell

(a) (b) FhGALE M U I S W1 W2 E1 E2 E3 M - +

Keratinocyte-Releasable Stratifin Functions as a Potent Collagenase-Stimulating Factor in Fibroblasts Aziz Ghahary, Feridoun Karimi-Busheri, Yvonne Marcoux,

The Nuclear Hat1p/Hat2p Complex

Selective Degradation of Ubiquitinated Sic1 by Purified 26S Proteasome Yields Active S Phase Cyclin-Cdk Rati Verma, Hayes McDonald, John R Yates, Raymond.

Volume 48, Issue 2, Pages (October 2005)

The Intrinsically Disordered Sem1 Protein Functions as a Molecular Tether during Proteasome Lid Biogenesis Robert J. Tomko, Mark Hochstrasser Molecular.

Class C Vps Protein Complex Regulates Vacuolar SNARE Pairing and Is Required for Vesicle Docking/Fusion Trey K. Sato, Peter Rehling, Michael R. Peterson,

Volume 96, Issue 5, Pages (March 1999)

Purification of chromogranin B from over-expressing insect sf9 cells.

Distinct Pores for Peroxisomal Import of PTS1 and PTS2 Proteins

Fus3-Regulated Tec1 Degradation through SCFCdc4 Determines MAPK Signaling Specificity during Mating in Yeast Song Chou, Lan Huang, Haoping Liu Cell

Volume 38, Issue 5, Pages (June 2010)

Molecular Therapy - Methods & Clinical Development

The Mitochondrial Presequence Translocase

Volume 3, Issue 2, Pages (August 2002)

Volume 96, Issue 3, Pages (February 1999)

Volume 11, Issue 24, Pages (December 2001)

Dimitra J. Mitsiou, Hendrik G. Stunnenberg Molecular Cell

Md Nasimuzzaman, Danielle Lynn, Johannes CM van der Loo, Punam Malik

Multiubiquitin Chain Receptors Define a Layer of Substrate Selectivity in the Ubiquitin- Proteasome System Rati Verma, Robert Oania, Johannes Graumann,

Protein purification and quality control.

Volume 39, Issue 5, Pages (September 2010)

George Simos, Anke Sauer, Franco Fasiolo, Eduard C Hurt Molecular Cell

Michael J. Lee, Henrik G. Dohlman Current Biology

Identification of GRP78 and vaspin complex by tandem affinity tag purification. Identification of GRP78 and vaspin complex by tandem affinity tag purification.

Volume 89, Issue 4, Pages (May 1997)

An Activator Target in the RNA Polymerase II Holoenzyme

WW domain does not show dominant-negative effect on TBSV replication in a yeast cell extract. WW domain does not show dominant-negative effect on TBSV.

Mapping the Pirh2 and p73 interaction sites.

DNA-binding and double-strand break formation by Vpr.

SDS-PAGE and Coomassie Blue Staining of Protein Samples after Various Purification Steps of Both Rice and Barley ADPG Hydrolytic NPPs.Lane numbering of.

Acetylation Regulates Transcription Factor Activity at Multiple Levels

Presentation transcript:

His-eIF5A Lys Elution FTL Online Resource Figure 1. Purification of 6xHis-eIF5A Lys from E. coli. E. coli strain M15 was transformed with pQE-eIF5A and production of the recombinant yeast protein 6xHis-eIF5A Lys was obtained after induction of expression. The cell pellet was washed and suspended in ice-cold lysis buffer. The 6xHis-eIF5A Lys fusion protein was purified by affinity chromatography on Ni-NTA resin and fractions were analyzed by SDS-PAGE with Coomassie staining. Lanes 2 and 3 contain the flowthrough and the washing fractions. The 6xHis- eIF5A Lys was eluted in fractions 4 to 12. The purified protein from lanes 7 to 11 were used in Figure 1. eIF5A dimerizes not only in vitro but also in vivo and its molecular envelope is similar to the EF-P monomer Camila Arnaldo Olhê Dias a, Wanius Garcia b, Cleslei Fernando Zanelli a, Sandro Roberto Valentini a, * a Department of Biological Sciences, School of Pharmaceutical Sciences, UNESP – Univ Estadual Paulista, Araraquara, SP, Brazil b Center of Natural Sciences and Humanities, UFABC – Univ Federal do ABC, Santo André, SP, Brazil. * Corresponding autho: Faculdade de Ciências Farmacêuticas – UNESP, Rodovia Araraquara-Jaú, km 01, Araraquara, SP, Brazil, , Telephone: , Fax: , 12

Online Resource Figure 2. SDS-PAGE of eIF5A proteins purified from bacteria and yeast. The proteins purified by ion exchange (bacteria) or affinity chromatography (yeast) were analyzed by SDS-PAGE with Coomassie staining. Lane 2 shows the recombinant yeast eIF5A Hyp purified from bacteria. Lanes 3 and 4 contain the proteins 6xHis-eIF5A Hyp and 6xHis-eIF5A K51R purified from yeast. These proteins were used in Figures 1 and 2. His-eIF5A K51R purified from yeast eIF5A Hyp purified from bacteria His-eIF5A Hyp purified from yeast

Online Resource Figure 3. Absorption spectra of purified proteins. (a) Recombinant yeast 6xHis-eIF5A Lys protein purified from bacteria and used in Figure 1. (b) Recombinant yeast eIF5A Hyp protein purified from bacteria and used in Figure 1. (c) Recombinant yeast eIF5A Lys protein purified from bacteria for the GST- pulldown in vitro assay (Figure 3c). a b c