Journal: Oncotarget

Article Title: LINE-1 hypomethylation in normal colon mucosa is associated with poor survival in Chinese patients with sporadic colon cancer

doi:

Figure Lengend Snippet: Bisulfite-treated DNA samples from adjacent normal mucosa were subjected to PCR amplification and were quantitatively analyzed by pyrosequencing. The C base marked in yellow served as a quality control of the bisulfite conversion efficiency. Four analyzed CpG sites are highlighted in blue, and the percent methylation rate is provided for each site. The mean percentage was computed as the LINE-1 methylation rate (LMR) for each case. Two cases with relatively higher (73.8%, A. ) or lower (19.4%, B. ) LMR were shown, respectively.

Article Snippet: In this system, a bisulfite-converted universal human DNA standard of 100% methylation (#D5015, ZYMO Research, USA) and ALU-C4 were used as the reference template and internal control, respectively.

Techniques: Amplification, Methylation

Journal: American Journal of Physiology - Cell Physiology

Article Title: miR-222 represses expression of zipcode binding protein-1 and phospholipase C-γ1 in intestinal epithelial cells

doi: 10.1152/ajpcell.00165.2018

Figure Lengend Snippet: Interaction of microRNA-222 (miR-222) with the zipcode binding protein-1 (ZBP1) and phospholipase C-γ1 (PLCγ1) mRNAs. A: miR-222 and U6 RNA levels in HCT-116 cells transfected with biotinylated miR-222 for 24 h. Values are means ± SE from 3 independent experiments (n = 4). *P < 0.01 compared with cells transfected with control scramble oligomer as analyzed by one-way ANOVA followed by Duncan’s test. B: levels of ZBP1, PLCγ1, claudin-1 (CCND1), and FZD7 mRNAs in the materials pulled down by biotin-miR-222 (left) and total input mRNAs (right) in cells described in A. Fzd7 served as a positive control.

Article Snippet: Human ZBP1 and PLCγ1 cDNAs and siRNAs were purchased from OriGene Technologies (Rockville, MD).

Techniques: Binding Assay, Transfection, Positive Control

Journal: American Journal of Physiology - Cell Physiology

Article Title: miR-222 represses expression of zipcode binding protein-1 and phospholipase C-γ1 in intestinal epithelial cells

doi: 10.1152/ajpcell.00165.2018

Figure Lengend Snippet: Ectopically expressed microRNA-222 (miR-222) represses the expression of zipcode binding protein-1 (ZBP1) and phospholipase C-γ1 (PLCγ1). A: levels of miR-222 and U6 RNA 48 h after transfection with pre-miR-222 as measured by quantitative PCR analysis. Values are means ± SE from independent experiments (n = 4). *P < 0.05, compared with cells transfected with control scrambled oligomer analyzed by one-way ANOVA followed by Duncan’s test. B: immunoblots of ZBP1, PLCγ1, and PCNA proteins in HCT-116 cells described in A. Whole cell lysates were harvested and prepared for Western blotting; equal loading was monitored by assessing GAPDH levels. C: quantitative analysis of ZBP1 and PLCγ1 immunoblotting signals as measured by densitometry using Bio-Rad-XRS system equipped with Image laboratory software (version 4.1) and used “Quantity tool” to determine the band intensity volume. The values were normalized with internal loading control GAPDH. Values are means ± SE of data from 3 independent experiments (n = 3). D and E: changes in ZBP1, PLCγ1, and PCNA proteins in IEC-Cdx2L1 cells 48 h after transfection with pre-miR-222. Values are means ± SE (n = 3). Statistical test: means are compared with the scramble (cells exposed to pre-miR-222) by nonparametric comparison (*P < 0.0495, Kruskal-Wallis test).

Article Snippet: Human ZBP1 and PLCγ1 cDNAs and siRNAs were purchased from OriGene Technologies (Rockville, MD).

Techniques: Expressing, Binding Assay, Transfection, Real-time Polymerase Chain Reaction, Western Blot, Software

Journal: American Journal of Physiology - Cell Physiology

Article Title: miR-222 represses expression of zipcode binding protein-1 and phospholipase C-γ1 in intestinal epithelial cells

doi: 10.1152/ajpcell.00165.2018

Figure Lengend Snippet: microRNA-222 (miR-222) overexpression destabilizes the zipcode binding protein-1 (ZBP1) and phospholipase C-γ1 (PLCγ1) mRNAs. A: levels of the ZBP1 and PLCγ1 mRNAs in cells transfected with pre-miR-222 for 48 h. JunD served as a negative control. Values are the means ± SE from independent experiments (n = 3). *P < 0.01, compared with cells transfected with control scramble oligomer as analyzed by one-way ANOVA followed by Duncan’s test. B–D: half-lives of the ZBP1, PLCγ1, and GAPDH mRNA in cells described in A. Total cellular RNA was isolated at indicated times after administration of actinomycin D (5 μg/ml), and the levels of ZBP1, PLCγ1, and GAPDH mRNAs were measured by quantitative PCR analysis. GAPDH mRNA served as a control. *P < 0.05, compared with cells transfected with C-oligo as analyzed by one-way ANOVA followed by Duncan’s test.

Article Snippet: Human ZBP1 and PLCγ1 cDNAs and siRNAs were purchased from OriGene Technologies (Rockville, MD).

Techniques: Over Expression, Binding Assay, Transfection, Negative Control, Isolation, Real-time Polymerase Chain Reaction

Journal: American Journal of Physiology - Cell Physiology

Article Title: miR-222 represses expression of zipcode binding protein-1 and phospholipase C-γ1 in intestinal epithelial cells

doi: 10.1152/ajpcell.00165.2018

Figure Lengend Snippet: microRNA-222 (miR-222) silencing enhances the expression of zipcode binding protein-1 (ZBP1) and phospholipase C-γ1 (PLCγ1). A: levels of miR-222 and U6 RNA 48 h after transfection with anti-miR-222. Values are the means ± SE from 3 independent experiments (n = 4). *P < 0.05, compared with cells transfected with control oligomer (C-oligo). B: immunoblots of ZBP1, PLCγ, and PCNA proteins in cells described in A. C: quantitative analysis of ZBP1 and PLCγ1 immunoblotting signals by densitometry using Bio-Rad-XRS system equipped with Image laboratory software (version 4.1) and used “Quantity tool” to determine the band intensity volume. The values were normalized with internal loading control GAPDH. Values are means ± SE of data from 3 independent experiments (n = 3). Statistical test: means are compared with the scramble (cells exposed to Anti-miR-222) by nonparametric comparison (*P < 0.0495, Kruskal-Wallis test).

Article Snippet: Human ZBP1 and PLCγ1 cDNAs and siRNAs were purchased from OriGene Technologies (Rockville, MD).

Techniques: Expressing, Binding Assay, Transfection, Western Blot, Software

Journal: American Journal of Physiology - Cell Physiology

Article Title: miR-222 represses expression of zipcode binding protein-1 and phospholipase C-γ1 in intestinal epithelial cells

doi: 10.1152/ajpcell.00165.2018

Figure Lengend Snippet: microRNA-222 (miR-222) silencing increases the stability of the zipcode binding protein-1 (ZBP1) and phospholipase C-γ1 (PLCγ1) mRNAs. A: levels of the ZBP1 and PLCγ1 mRNAs in cells transfected with anti-miR-222 for 48 h. JunD served as a negative control. Values are the means ± SE from independent experiments (n = 4). *P < 0.01, compared with cells transfected with C-oligo. B–D: half-lives of the ZBP1, PLCγ1, and GAPDH mRNA in cells described in A. Total cellular RNA was isolated at indicated times after administration of actinomycin D (5 μg/ml), and the levels of ZBP1, PLCγ1, and GAPDH mRNAs were measured by quantitative PCR analysis. GAPDH mRNA served as a control. *P < 0.05, compared with cells transfected with C-oligo as analyzed by one-way ANOVA followed by Duncan’s test.

Article Snippet: Human ZBP1 and PLCγ1 cDNAs and siRNAs were purchased from OriGene Technologies (Rockville, MD).

Techniques: Binding Assay, Transfection, Negative Control, Isolation, Real-time Polymerase Chain Reaction

Journal: American Journal of Physiology - Cell Physiology

Article Title: miR-222 represses expression of zipcode binding protein-1 and phospholipase C-γ1 in intestinal epithelial cells

doi: 10.1152/ajpcell.00165.2018

Figure Lengend Snippet: microRNA-222 (miR-222)-regulated expression of zipcode binding protein-1 (ZBP1) and phospholipase C-γ1 (PLCγ1) modulates rapid epithelial restitution after wounding. A: images of cell migration: a, 0 h after wounding in control cells (Con-0 h); b, 6 h after wounding in control cells (Con-6 h); c, 6 h after wounding in cells transfected with scramble oligo (C-oligo-6 h); d, 6 h after wounding in cells transfected with pre-miR-222 alone for 48 h (Pre-miR-222–6 h); e, 6 h after wounding in cells cotransfected with pre-miR-222 and the expression vector encoding ZBP1 (Pre-miR222 + ZBP1); f, 6 h after wounding in cells cotransfected with pre-miR-222 and the expression vector encoding PLCγ1 (Pre-miR222+ PLCγ1). Scale bar, 100 μm; magnification, ×100. B: summarized data showing rates of cell migration 6 h after wounding in cells described in A. Values are the means ± SE of data from 6 dishes and repeated 4 times independently (n = 4). *,+P < 0.05, compared with cells transfected with scramble and cells transfected with pre-miR-222, respectively as analyzed by one-way ANOVA followed by Duncan’s test. C: images of cell migration: a: 0 h after wounding in control cells; b: 6 h after wounding in control cells; c, 6 h after wounding in cells transfected with scramble oligo; d: 6 h after wounding in cells transfected with anti-miR-222 alone for 48 h; e: 6 h after wounding in cells cotransfected with anti-miR-222 and siZBP1 (Anti-miR222 + siZBP1); f: 6 h after wounding in cells cotransfected with anti-miR-222 and siPLCγ1 (Anti-miR222 + siPLCγ1); and g: 6 h after wounding in cells cotransfected with anti-miR-222, siZBP1 and siPLCγ1 (anti-miR222 + siZBP1 + siPLCγ1). Scale bar, 100 μm; magnification, ×100. D: summarized data showing rates of cell migration in cells described in C. Values are the means ± SE of data from 6 dishes and repeated four times independently (n = 4). *,+P < 0.05, compared with cells transfected with scramble and cells transfected with Anti-miR-222, respectively as analyzed by one-way ANOVA followed by Duncan’s test. E: immunoblot of PCNA protein in nonwounding and 0 h and 6 h after wounding (#1 and #2). Whole cell lysates were harvested and prepared for Western blotting; equal loading was monitored by assessing GAPDH levels.

Article Snippet: Human ZBP1 and PLCγ1 cDNAs and siRNAs were purchased from OriGene Technologies (Rockville, MD).

Techniques: Expressing, Binding Assay, Migration, Transfection, Plasmid Preparation, Western Blot

Journal: Scientific Reports

Article Title: New Insights into Cooperative Binding of Homeodomain Transcription Factors PREP1 and PBX1 to DNA

doi: 10.1038/srep40665

Figure Lengend Snippet: Panel A: Homeodomain (HD) protein constructs used for characterizations described in this study, in comparison with full-length proteins. PBX1 HD and HOXB1 HD constructs are the same used in the X-ray crystallographic study . PREP1 HD is present in four lengths, PREP1 257-325 (PREP1 hd ), PREP1 240-344 (PREP1 HD ) and two intermediate lengths (PREP1 257-344 - PREP1 hd-C - and PREP1 240-325 -PREP1 hd-N -), where respectively the N- or C-terminal extensions of the HD are omitted. Production of the proteins is described in the Materials and Methods section. PBC-A and PBC-B domains of PBX1 are those required for its dimerization with PREP1. TALE lies between helix 1 and helix 2 of the HD. MEIS-A and MEIS-B domains are PREP1 motifs required for heterodomerization with PBX1. Panel B: Consensus sequences for DNA binding by PREP1-PBX1: the very frequent decameric (PMH) and the less frequent octameric (PH) oligonucleotides , are both highlighted in red in the sequence. Control probes correspond to two sequences that do not contain any PBX1 orPREP1 binding site. Oligonucleotides used for FP were 5′ labelled with 6-carboxyfluorescein (6-FAM) (see the Materials and Methods).

Article Snippet: The DNA template of PREP1 HD in which the four residues K 231 , K 233 , K 234 , and K 235 were mutated to alanine was purchased from Genscript Corporation (Piscataway, NJ) and showed the following sequence: 5′CAGCTTCAGTTACAGTTAAACCAAGATCTCAGCATCTTGCATCAAGATGATGGTTCATCTAAGAACAAGAGGGGCGTCCTGCCAAAGCATGCCACGAACGTGATGCGGTCCTGGCTCTTCCAGCACATCGGGCATCCCTACCCAACAGAGGATGAGAAAAAACAGATTGCTGCTCAGACAAATTTGACACTACTCCAAGTCAACAACTGGTTCATCAATGCCAGAAGACGAATTCTTCAGCCAATGTTGGATTCAAGTTGTTCAGAGACCCCCGCAACAGCGGCAGCAACTGCTCAGAACCGGCCAGTTCAGAGG 3′.

Techniques: Construct, Comparison, Binding Assay, Sequencing, Control

Journal: Scientific Reports

Article Title: New Insights into Cooperative Binding of Homeodomain Transcription Factors PREP1 and PBX1 to DNA

doi: 10.1038/srep40665

Figure Lengend Snippet: K D values for individual HDs with different DNA sequences, measured by FP.

Article Snippet: The DNA template of PREP1 HD in which the four residues K 231 , K 233 , K 234 , and K 235 were mutated to alanine was purchased from Genscript Corporation (Piscataway, NJ) and showed the following sequence: 5′CAGCTTCAGTTACAGTTAAACCAAGATCTCAGCATCTTGCATCAAGATGATGGTTCATCTAAGAACAAGAGGGGCGTCCTGCCAAAGCATGCCACGAACGTGATGCGGTCCTGGCTCTTCCAGCACATCGGGCATCCCTACCCAACAGAGGATGAGAAAAAACAGATTGCTGCTCAGACAAATTTGACACTACTCCAAGTCAACAACTGGTTCATCAATGCCAGAAGACGAATTCTTCAGCCAATGTTGGATTCAAGTTGTTCAGAGACCCCCGCAACAGCGGCAGCAACTGCTCAGAACCGGCCAGTTCAGAGG 3′.

Techniques: Control

Journal: Scientific Reports

Article Title: New Insights into Cooperative Binding of Homeodomain Transcription Factors PREP1 and PBX1 to DNA

doi: 10.1038/srep40665

Figure Lengend Snippet: DNA probes were incubated with different amounts of HD. Protein:DNA ratios were 0.5 (DNA excess), 1 (equimolar ratio), or 2 (protein excess). Gels were stained both with ethidium bromide and coomassie blue to visualize DNA or proteins, respectively. Panel A: PREP1 HD forms two different complexes with PMH: the first is visible, at protein:DNA ratio of 0.5 (lanes 1 and 2, left panel), and the second as a smeared band, at protein:DNA ratio of 2 (lane 3, left panel). This slower migrating band might represent binding of two PREP1 HD proteins to DNA. In the case of PH (central panel) and control probes (right panel), a specific DNA complex is not formed. Panel B: PREP1 hd forms a single complex with PMH; whereas binding to control and PH oligos is weak and no specific bands are formed. Panel C: PBX1 HD forms a single complex with PMH and PH oligos at protein:DNA ratio of 0.5 (lane 1); by increasing the protein concentration, a slower migrating band becomes visible, possibly due to binding of a second monomer to DNA. PBX1 binds also the control oligo.

Article Snippet: The DNA template of PREP1 HD in which the four residues K 231 , K 233 , K 234 , and K 235 were mutated to alanine was purchased from Genscript Corporation (Piscataway, NJ) and showed the following sequence: 5′CAGCTTCAGTTACAGTTAAACCAAGATCTCAGCATCTTGCATCAAGATGATGGTTCATCTAAGAACAAGAGGGGCGTCCTGCCAAAGCATGCCACGAACGTGATGCGGTCCTGGCTCTTCCAGCACATCGGGCATCCCTACCCAACAGAGGATGAGAAAAAACAGATTGCTGCTCAGACAAATTTGACACTACTCCAAGTCAACAACTGGTTCATCAATGCCAGAAGACGAATTCTTCAGCCAATGTTGGATTCAAGTTGTTCAGAGACCCCCGCAACAGCGGCAGCAACTGCTCAGAACCGGCCAGTTCAGAGG 3′.

Techniques: Incubation, Staining, Binding Assay, Control, Protein Concentration

Journal: Scientific Reports

Article Title: New Insights into Cooperative Binding of Homeodomain Transcription Factors PREP1 and PBX1 to DNA

doi: 10.1038/srep40665

Figure Lengend Snippet: FP K D measurement of PREP1 HDs in the presence of a preformed PBX1 HD : DNA complex.

Article Snippet: The DNA template of PREP1 HD in which the four residues K 231 , K 233 , K 234 , and K 235 were mutated to alanine was purchased from Genscript Corporation (Piscataway, NJ) and showed the following sequence: 5′CAGCTTCAGTTACAGTTAAACCAAGATCTCAGCATCTTGCATCAAGATGATGGTTCATCTAAGAACAAGAGGGGCGTCCTGCCAAAGCATGCCACGAACGTGATGCGGTCCTGGCTCTTCCAGCACATCGGGCATCCCTACCCAACAGAGGATGAGAAAAAACAGATTGCTGCTCAGACAAATTTGACACTACTCCAAGTCAACAACTGGTTCATCAATGCCAGAAGACGAATTCTTCAGCCAATGTTGGATTCAAGTTGTTCAGAGACCCCCGCAACAGCGGCAGCAACTGCTCAGAACCGGCCAGTTCAGAGG 3′.

Techniques:

Journal: Scientific Reports

Article Title: New Insights into Cooperative Binding of Homeodomain Transcription Factors PREP1 and PBX1 to DNA

doi: 10.1038/srep40665

Figure Lengend Snippet: Panel A: A sample containing a preformed PBX1 HD :PMH complex (at 4 and 8 μM, respectively) was titrated with increasing amounts of PREP1 HD (2, 4 and 8 μM). PREP1 HD binds preferentially to the PBX1 HD :PMH complex rather than to the free DNA; the DNA pool decrease appreciably upon an increase in PREP1 HD concentration. At the lower PREP1 HD concentrations (lane 3) a slower migrating band is observed above the monomeric PREP1 HD :DNA and PBX1 HD :DNA complexes (identified in lanes 1 and 2, respectively). The band marked with a star in lane 5 was analyzed by mass spectrometry. Panel B: Titration of PREP1 HD : PMH complex with PBX1 HD . PREP1 HD (4 μM) and PMH (4 μM) were titrated with increasing concentrations of PBX1 hHD (2, 4 and 8 μM). Upon increase of PBX1 HD a slower migrating band appears, corresponding to PREP1 HD :DNA:PBX1 HD complex. Panel C: Titration of PBX1 HD : PMH complex with PREP1 hd . PBX1 HD (4 μM) and PMH (4 μM) were titrated with increasing concentrations of PREP1 hd (2, 4 and 8 μM). Upon increase of PREP1 hd only a band corresponding to PREP1 hd :DNA is visible, therefore, PREP1 hd in EMSA does not show clear binding to the preformed PBX1 HD :DNA complex. Panel D: Titration of PMH:PBX1 HD with PREP1 hd-N . PMH and PBX1 HD at a fixed concentration (8 μM and 4 μM, respectively) were titrated with increasing amounts of PREP1 hd-N (2, 4, 8, and 16 μM). Upon an increase in PREP1 hd-N no retarded band above the individual PMH:PREP1 hd-N and PMH:PBX11 HD complexes (lanes 3-5) are observed Panel E: Titration of PBX1 HD :PMH with PREP1 hd-C . PMH oligo and PBX1 HD at a fixed concentration (8 μM and 4 μM, respectively) were titrated with increasing concentrations of PREP1 hd-C (2, 4 and 8 μM). Upon an increase in PREP1 hd - C , a slower migrating band corresponding to PREP1 hd-C :DNA: PBX1 HD complex is visible.

Article Snippet: The DNA template of PREP1 HD in which the four residues K 231 , K 233 , K 234 , and K 235 were mutated to alanine was purchased from Genscript Corporation (Piscataway, NJ) and showed the following sequence: 5′CAGCTTCAGTTACAGTTAAACCAAGATCTCAGCATCTTGCATCAAGATGATGGTTCATCTAAGAACAAGAGGGGCGTCCTGCCAAAGCATGCCACGAACGTGATGCGGTCCTGGCTCTTCCAGCACATCGGGCATCCCTACCCAACAGAGGATGAGAAAAAACAGATTGCTGCTCAGACAAATTTGACACTACTCCAAGTCAACAACTGGTTCATCAATGCCAGAAGACGAATTCTTCAGCCAATGTTGGATTCAAGTTGTTCAGAGACCCCCGCAACAGCGGCAGCAACTGCTCAGAACCGGCCAGTTCAGAGG 3′.

Techniques: Concentration Assay, Mass Spectrometry, Titration, Binding Assay

Journal: Scientific Reports

Article Title: New Insights into Cooperative Binding of Homeodomain Transcription Factors PREP1 and PBX1 to DNA

doi: 10.1038/srep40665

Figure Lengend Snippet: Mass spectrometry analysis of EMSA titrations bands from Fig. 3 .

Article Snippet: The DNA template of PREP1 HD in which the four residues K 231 , K 233 , K 234 , and K 235 were mutated to alanine was purchased from Genscript Corporation (Piscataway, NJ) and showed the following sequence: 5′CAGCTTCAGTTACAGTTAAACCAAGATCTCAGCATCTTGCATCAAGATGATGGTTCATCTAAGAACAAGAGGGGCGTCCTGCCAAAGCATGCCACGAACGTGATGCGGTCCTGGCTCTTCCAGCACATCGGGCATCCCTACCCAACAGAGGATGAGAAAAAACAGATTGCTGCTCAGACAAATTTGACACTACTCCAAGTCAACAACTGGTTCATCAATGCCAGAAGACGAATTCTTCAGCCAATGTTGGATTCAAGTTGTTCAGAGACCCCCGCAACAGCGGCAGCAACTGCTCAGAACCGGCCAGTTCAGAGG 3′.

Techniques: Mass Spectrometry

Journal: Scientific Reports

Article Title: New Insights into Cooperative Binding of Homeodomain Transcription Factors PREP1 and PBX1 to DNA

doi: 10.1038/srep40665

Figure Lengend Snippet: Panel A: Superposition of 1 H- 15 N HSQC spectra of PREP1 HD (magenta) and PREP1 hd (blue). Panel B: Combined amide chemical shift difference (Δ δ ) between PREP1 HD and PREP1 hd corresponding residues. Panel C: Cartoon representation of the PREP1 HD model: the residues (in blue) shared with PREP1 hd , the N- and C- terminal extensions (in cyan) of PREP1 HD . Spheres indicate PREP1 HD residues showing significant (>average) amide Δ δ with respect to PREP1 hd . Panel D: Normalized CD melting curves for PREP1 HD (magenta) and PREP1 hd (blue) at 222 nm.

Article Snippet: The DNA template of PREP1 HD in which the four residues K 231 , K 233 , K 234 , and K 235 were mutated to alanine was purchased from Genscript Corporation (Piscataway, NJ) and showed the following sequence: 5′CAGCTTCAGTTACAGTTAAACCAAGATCTCAGCATCTTGCATCAAGATGATGGTTCATCTAAGAACAAGAGGGGCGTCCTGCCAAAGCATGCCACGAACGTGATGCGGTCCTGGCTCTTCCAGCACATCGGGCATCCCTACCCAACAGAGGATGAGAAAAAACAGATTGCTGCTCAGACAAATTTGACACTACTCCAAGTCAACAACTGGTTCATCAATGCCAGAAGACGAATTCTTCAGCCAATGTTGGATTCAAGTTGTTCAGAGACCCCCGCAACAGCGGCAGCAACTGCTCAGAACCGGCCAGTTCAGAGG 3′.

Techniques:

Journal: Scientific Reports

Article Title: New Insights into Cooperative Binding of Homeodomain Transcription Factors PREP1 and PBX1 to DNA

doi: 10.1038/srep40665

Figure Lengend Snippet: Data reported correspond to a protein:PMH molar ratio of 1:0.5. Panel A: Superposition of 1 H- 15 N HSQC spectra of free (black) and PMH-bound (red) PREP1 HD . (left) and PREP1 hd (right) Panel B: Plot showing the peaks intensity reduction observed upon PMH binding to PREP1 hd and PREP1 HD . Panel C: Residues disappearing or showing significant amide chemical shift displacement (as reported in the CSD plot in ) are highlighted on PREP1 HD structural model (top, generated using CS23D2.0 web server) and PREP1 hd NMR structure (bottom, 1 × 2 N.pdb,).

Article Snippet: The DNA template of PREP1 HD in which the four residues K 231 , K 233 , K 234 , and K 235 were mutated to alanine was purchased from Genscript Corporation (Piscataway, NJ) and showed the following sequence: 5′CAGCTTCAGTTACAGTTAAACCAAGATCTCAGCATCTTGCATCAAGATGATGGTTCATCTAAGAACAAGAGGGGCGTCCTGCCAAAGCATGCCACGAACGTGATGCGGTCCTGGCTCTTCCAGCACATCGGGCATCCCTACCCAACAGAGGATGAGAAAAAACAGATTGCTGCTCAGACAAATTTGACACTACTCCAAGTCAACAACTGGTTCATCAATGCCAGAAGACGAATTCTTCAGCCAATGTTGGATTCAAGTTGTTCAGAGACCCCCGCAACAGCGGCAGCAACTGCTCAGAACCGGCCAGTTCAGAGG 3′.

Techniques: Binding Assay, Generated