scholarly journals The N-terminal zinc finger domain of Tgf2 transposase contributes to DNA binding and to transposition activity

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Xia-Yun Jiang ◽  
Fei Hou ◽  
Xiao-Dan Shen ◽  
Xue-Di Du ◽  
Hai-Li Xu ◽  
...  
Keyword(s):  
Dna Binding ◽  
Zinc Finger ◽  
Zinc Finger Domain ◽  
Transposition Activity

Characterization of the DNA Binding Activity of the ZFY Zinc Finger Domain

Biochemistry ◽  
2010 ◽  
Vol 49 (4) ◽  
pp. 679-686 ◽  
Author(s):  
Jennifer Grants ◽  
Erin Flanagan ◽  
Andrea Yee ◽  
Paul J. Romaniuk
Keyword(s):  
Dna Binding ◽  
Zinc Finger ◽  
Binding Activity ◽  
Zinc Finger Domain ◽  
Dna Binding Activity

P1608Inhibition of GATA4 dimerization suppress hypertrophic responses

2019 ◽  
Vol 40 (Supplement_1) ◽  
Author(s):  
S Shimizu ◽  
Y Sunagawa ◽  
K Hara ◽  
A Hishiki ◽  
Y Katanasaka ◽  
...  
Keyword(s):  
Heart Failure ◽  
Dna Binding ◽  
Cardiac Hypertrophy ◽  
Zinc Finger ◽  
Therapeutic Target ◽  
Transcriptional Activity ◽  
Luciferase Assay ◽  
Zinc Finger Motif ◽  
Zinc Finger Domain ◽  
Acetylation Site

Abstract Introduction Hypertrophic signals eventually reach the nuclei of cardiomyocytes, change patterns of gene expression, and cause the development of heart failure. During the development of heart failure, intrinsic histone acetyltransferase called p300 induce GATA4 acetylation. Acetylated GATA4 increases its DNA binding, up-regulates cardiac hypertrophic response genes, and lead to heart failure. A zinc finger protein, GATA4 is the transcription factor that expression level is high in heart. It has been reported that GATA1, the same GATA family, regulates transcriptional activity through its homo-dimerization. However, GATA4 homo-dimerization and its relationship to hypertrophic responses are still unknown. Purpose To clarify the relationship between GATA4 homo-dimerization and transcriptional activity and investigate whether inhibition of this homo-dimerization become therapeutic target for cardiac hypertrophy. Methods GST pull-down and DNA pull-down assay were performed using GST fusion full length and deletion mutants of GATA4 and biotin-conjugated ET-1 promoter probe including a GATA element. Recombinant C-zinc finger domain (256–326), including C-zinc finger motif (256–295) and acetylation site (308–326) was cross-linked using glutaraldehyde and subjected to silver staining. An expression plasmid with three GATA4-acetylation site mutant-conjugated with nuclear localization sequence (3xG4D) was constructed. Immunoprecipitation and western blotting were performed using nuclear extract from HEK293T cells expressing p300, GATA4, and 3xG4D. Luciferase assay was using ANF and ET-1 promoter sequences. Neonatal rat cultured cardiomyocyte expressed 3xG4D and then stimulated with phenylephrine (PE) for 48 hours. Next cardiomyocytes stained with α-actinin antibody and measured the cell surface area. Results The acetylation site of GATA4 was required for the dimerization of GATA4. But, C-zinc finger motif (256–295) and the acetylation site were required for the DNA binding. Recombinant C-zinc finger domain formed not only a homo-dimer but also a multimer. Co-expression of p300 increased the formation of homo-dimer as well as the acetylation of GATA4 in HEK293T cells. The GATA4 homo-dimer was disrupted by acetyl-deficient GATA4 or HAT-deficient p300 mutant. Overexpression of 3xG4D prevented the dimerization of GATA4, but not acetylation of GATA4. The result of luciferase assay showed that overexpression of 3xG4D prevented p300/GATA-induced ANF and ET-1 promoter activities. Furthermore, overexpression of 3xG4D inhibited phenylephrine-induced cardiomyocyte hypertrophy. Conclusions These results suggest that GATA4 dimerization may play an important role in hypertrophy-response gene activation. Thus, it is likely that inhabitation of GATA4 dimerization become therapeutic target for cardiac hypertrophy.

scholarly journals The structural role of the zinc ion can be dispensable in prokaryotic zinc-finger domains

2009 ◽  
Vol 106 (17) ◽  
pp. 6933-6938 ◽  
Author(s):  
Ilaria Baglivo ◽  
Luigi Russo ◽  
Sabrina Esposito ◽  
Gaetano Malgieri ◽  
Mario Renda ◽  
...  
Keyword(s):  
Dna Binding ◽  
Zinc Finger ◽  
Coordination Sphere ◽  
Zinc Ion ◽  
Binding Domain ◽  
Hydrophobic Core ◽  
Zinc Finger Domain ◽  
High Sequence Identity ◽  
Zinc Coordination ◽  
Zinc Finger Domains

The recent characterization of the prokaryotic Cys2His2 zinc-finger domain, identified in Ros protein from Agrobacterium tumefaciens, has demonstrated that, although possessing a similar zinc coordination sphere, this domain is structurally very different from its eukaryotic counterpart. A search in the databases has identified ≈300 homologues with a high sequence identity to the Ros protein, including the amino acids that form the extensive hydrophobic core in Ros. Surprisingly, the Cys2His2 zinc coordination sphere is generally poorly conserved in the Ros homologues, raising the question of whether the zinc ion is always preserved in these proteins. Here, we present a functional and structural study of a point mutant of Ros protein, Ros56–142C82D, in which the second coordinating cysteine is replaced by an aspartate, 5 previously-uncharacterized representative Ros homologues from Mesorhizobium loti, and 2 mutants of the homologues. Our results indicate that the prokaryotic zinc-finger domain, which in Ros protein tetrahedrally coordinates Zn(II) through the typical Cys2His2 coordination, in Ros homologues can either exploit a CysAspHis2 coordination sphere, previously never described in DNA binding zinc finger domains to our knowledge, or lose the metal, while still preserving the DNA-binding activity. We demonstrate that this class of prokaryotic zinc-finger domains is structurally very adaptable, and surprisingly single mutations can transform a zinc-binding domain into a nonzinc-binding domain and vice versa, without affecting the DNA-binding ability. In light of our findings an evolutionary link between the prokaryotic and eukaryotic zinc-finger domains, based on bacteria-to-eukaryota horizontal gene transfer, is discussed.

scholarly journals Cover Feature: Comparative Study of Photoswitchable Zinc‐Finger Domain and AT‐Hook Motif for Light‐Controlled Peptide–DNA Binding (Chem. Eur. J. 19/2019)

2019 ◽  
Vol 25 (19) ◽  
pp. 4866-4866
Author(s):  
Gosia M. Murawska ◽  
Claudia Poloni ◽  
Nadja A. Simeth ◽  
Wiktor Szymanski ◽  
Ben L. Feringa
Keyword(s):  
Dna Binding ◽  
Comparative Study ◽  
Zinc Finger ◽  
Zinc Finger Domain

scholarly journals In vitro analysis of the zinc-finger motif in human replication protein A

1999 ◽  
Vol 337 (2) ◽  
pp. 311-317 ◽  
Author(s):  
Jiaowang DONG ◽  
Jang-Su PARK ◽  
Suk-Hee LEE
Keyword(s):  
Dna Binding ◽  
Zinc Finger ◽  
Dna Polymerase ◽  
Protein A ◽  
Replication Protein A ◽  
Binding Domain ◽  
Replication Protein ◽  
Zinc Finger Motif ◽  
Zinc Finger Domain ◽  
Single Stranded Dna

Human replication protein A (RPA) is composed of 70, 34 and 11 kDa subunits (p70, p34 and p11 respectively) and functions in all three major DNA metabolic processes: replication, repair and recombination. Recent deletion analysis demonstrated that the large subunit of RPA, p70, has multiple functional domains, including a DNA polymerase α-stimulation domain and a single-stranded DNA-binding domain. It also contains a putative metal-binding domain of the 4-cysteine type (Cys-Xaa4-Cys-Xaa13-Cys-Xaa2-Cys) that is highly conserved among eukaryotes. To study the role of this domain in DNA metabolism, we created various p70 mutants that lack the zinc-finger motif (by Cys → Ala substitutions). Mutation at the zinc-finger domain (ZFM) abolished RPA's function in nucleotide excision repair (NER), but had very little impact on DNA replication. The failure of zinc-finger mutant RPA in NER may be explained by the observation that wild-type RPA significantly stimulated DNA polymerase δ activity, whereas only marginal stimulation was observed with zinc-finger mutant RPA. We also observed that ZFM reduced RPA's single-stranded DNA-binding activity by 2–3-fold in the presence of low amounts of RPA. Interestingly, the ZFM abolished phosphorylation of the p34 subunit by DNA-dependent protein kinase, but not that by cyclin-dependent kinase. Taker together, our results strongly suggest a positive role for RPA's zinc finger domain in its function.

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