Precise Coating of a Wide Range of DNA Templates by a Protein Polymer with a DNA Binding Domain

ACS Nano ◽  
2016 ◽  
Vol 11 (1) ◽  
pp. 144-152 ◽  
Author(s):  
Armando Hernandez-Garcia ◽  
Nicole A. Estrich ◽  
Marc W. T. Werten ◽  
Johan R. C. Van Der Maarel ◽  
Thomas H. LaBean ◽  
...  
Keyword(s):  
Dna Binding ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Dna Templates ◽  
Protein Polymer ◽  
Wide Range

scholarly journals Substitutions at Nonconserved Rheostat Positions Modulate Function by Rewiring Long-Range, Dynamic Interactions

2020 ◽  
Vol 38 (1) ◽  
pp. 201-214 ◽  
Author(s):  
Paul Campitelli ◽  
Liskin Swint-Kruse ◽  
S Banu Ozkan
Keyword(s):  
Dna Binding ◽  
Long Range ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Dynamic Coupling ◽  
Long Distance ◽  
Repressor Protein ◽  
Functional Changes ◽  
Lactose Repressor ◽  
Wide Range

Abstract Amino acid substitutions at nonconserved protein positions can have noncanonical and “long-distance” outcomes on protein function. Such outcomes might arise from changes in the internal protein communication network, which is often accompanied by changes in structural flexibility. To test this, we calculated flexibilities and dynamic coupling for positions in the linker region of the lactose repressor protein. This region contains nonconserved positions for which substitutions alter DNA-binding affinity. We first chose to study 11 substitutions at position 52. In computations, substitutions showed long-range effects on flexibilities of DNA-binding positions, and the degree of flexibility change correlated with experimentally measured changes in DNA binding. Substitutions also altered dynamic coupling to DNA-binding positions in a manner that captured other experimentally determined functional changes. Next, we broadened calculations to consider the dynamic coupling between 17 linker positions and the DNA-binding domain. Experimentally, these linker positions exhibited a wide range of substitution outcomes: Four conserved positions tolerated hardly any substitutions (“toggle”), ten nonconserved positions showed progressive changes from a range of substitutions (“rheostat”), and three nonconserved positions tolerated almost all substitutions (“neutral”). In computations with wild-type lactose repressor protein, the dynamic couplings between the DNA-binding domain and these linker positions showed varied degrees of asymmetry that correlated with the observed toggle/rheostat/neutral substitution outcomes. Thus, we propose that long-range and noncanonical substitutions outcomes at nonconserved positions arise from rewiring long-range communication among functionally important positions. Such calculations might enable predictions for substitution outcomes at a range of nonconserved positions.

scholarly journals Substitutions at Non-Conserved Rheostat Positions Modulate Function by Re-Wiring Long-Range, Dynamic Interactions

2020 ◽  
Author(s):  
Paul Campitelli ◽  
Liskin Swint-Kruse ◽  
S. Banu Ozkan
Keyword(s):  
Dna Binding ◽  
Long Range ◽  
Protein Function ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Dynamic Coupling ◽  
Long Distance ◽  
Modulate Function ◽  
Functional Changes ◽  
Wide Range

AbstractAmino acid substitutions at nonconserved protein positions can have non-canonical and “long-distance” outcomes on protein function. Such outcomes might arise from changes in the internal protein communication network, which is often accompanied by changes in structural flexibility. To test this, we calculated flexibilities (“DFI”) and dynamic coupling (“DCI”) for positions in the linker region of the lactose repressor protein (“LacI”). This region contains nonconserved positions for which substitutions alter DNA binding affinity. We first chose to study eleven substitutions at position 52. In computations, substitutions showed long-range effects on flexibilities of DNA binding positions, and the degree of flexibility change correlated with experimentally-measured changes in DNA binding. Substitutions also altered dynamic coupling to DNA binding positions in a manner that captured other experimentally-determined functional changes. Next, we broadened calculations to consider the dynamic coupling between 17 linker positions and the DNA binding domain. Experimentally, these linker positions exhibited a wide range of substitution outcomes: Four conserved positions tolerated almost no substitutions (“toggle”), ten nonconserved positions showed progressive changes from a range of substitutions (“rheostat”), and three nonconserved positions tolerated almost all substitutions (“neutral”). In computations with wild-type LacI, the dynamic couplings between the DNA binding domain and these linker positions showed varied degrees of asymmetry that correlated with the observed toggle/rheostat/neutral substitution outcomes. Thus, we propose that long-range and non-canonical substitutions outcomes at nonconserved positions arise from re-wiring long-range communication among functionally-important positions. Such calculations might enable predictions for substitution outcomes at a range of nonconserved positions.

scholarly journals A Quantitative Systems Approach to Define Novel Effects of Tumour p53 Mutations on Binding Oncoprotein MDM2

2021 ◽  
Vol 23 (1) ◽  
pp. 53
Author(s):  
Manuel Fuentes ◽  
Sanjeeva Srivastava ◽  
Angela M. Gronenborn ◽  
Joshua LaBaer
Keyword(s):  
Dna Binding ◽  
Protein Interactions ◽  
Hot Spot ◽  
Point Mutations ◽  
Protein Microarrays ◽  
Binding Domain ◽  
Structural Domain ◽  
Dna Binding Domain ◽  
Label Free ◽  
Wide Range

Understanding transient protein interactions biochemically at the proteome scale remains a long-standing challenge. Current tools developed to study protein interactions in high-throughput measure stable protein complexes and provide binary readouts; they do not elucidate dynamic and weak protein interactions in a proteome. The majority of protein interactions are transient and cover a wide range of affinities. Nucleic acid programmable protein arrays (NAPPA) are self-assembling protein microarrays produced by freshly translating full-length proteins in situ on the array surface. Herein, we have coupled NAPPA to surface plasmon resonance imaging (SPRi) to produce a novel label-free platform that measures many protein interactions in real-time allowing the determination of the KDs and rate constants. The developed novel NAPPA-SPRi technique showed excellent ability to study protein-protein interactions of clinical mutants of p53 with its regulator MDM2. Furthermore, this method was employed to identify mutant p53 proteins insensitive to the drug nutlin-3, currently in clinical practice, which usually disrupts the p53-MDM2 interactions. Thus, significant differences in the interactions were observed for p53 mutants on the DNA binding domain (Arg-273-Cys, Arg-273-His, Arg-248-Glu, Arg-280-Lys), on the structural domain (His-179-Tyr, Cys-176-Phe), on hydrophobic moieties in the DNA binding domain (Arg-280-Thr, Pro-151-Ser, Cys-176-Phe) and hot spot mutants (Gly-245-Cys, Arg-273-Leu, Arg-248-Glu, Arg-248-Gly), which signifies the importance of point mutations on the MDM2 interaction and nutlin3 effect, even in molecular locations related to other protein activities.

scholarly journals The Inhibition of Stat5 by a Peptide Aptamer Ligand Specific for the DNA Binding Domain Prevents Target Gene Transactivation and the Growth of Breast and Prostate Tumor Cells

2013 ◽  
Vol 6 (8) ◽  
pp. 960-987 ◽  
Author(s):  
Axel Weber ◽  
Corina Borghouts ◽  
Christian Brendel ◽  
Richard Moriggl ◽  
Natalia Delis ◽  
...  
Keyword(s):  
Dna Binding ◽  
Tumor Cells ◽  
Human Tumor ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Protein Activity ◽  
Intracellular Degradation ◽  
Wide Range ◽  
Peptide Aptamer ◽  
Prostate Tumor Cells

The signal transducer and activator of transcription Stat5 is transiently activated by growth factor and cytokine signals in normal cells, but its persistent activation has been observed in a wide range of human tumors. Aberrant Stat5 activity was initially observed in leukemias, but subsequently also found in carcinomas. We investigated the importance of Stat5 in human tumor cell lines. shRNA mediated downregulation of Stat5 revealed the dependence of prostate and breast cancer cells on the expression of this transcription factor. We extended these inhibition studies and derived a peptide aptamer (PA) ligand, which directly interacts with the DNA-binding domain of Stat5 in a yeast-two-hybrid screen. The Stat5 specific PA sequence is embedded in a thioredoxin (hTRX) scaffold protein. The resulting recombinant protein S5-DBD-PA was expressed in bacteria, purified and introduced into tumor cells by protein transduction. Alternatively, S5-DBD-PA was expressed in the tumor cells after infection with a S5-DBD-PA encoding gene transfer vector. Both strategies impaired the DNA-binding ability of Stat5, suppressed Stat5 dependent transactivation and caused its intracellular degradation. Our experiments describe a peptide based inhibitor of Stat5 protein activity which can serve as a lead for the development of a clinically useful compound for cancer treatment.

NMR studies of the R2 repeat and related peptide fragments of the DNA binding domain of c-Myb. New light on the structure and folding of R2.

1999 ◽  
Vol 96 (9/10) ◽  
pp. 1580-1584 ◽  
Author(s):  
I. Ségalas ◽  
S. Desjardins ◽  
H. Oulyadi ◽  
Y. Prigent ◽  
S. Tribouillard ◽  
...  
Keyword(s):  
Dna Binding ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Peptide Fragments ◽  
Nmr Studies ◽  
Related Peptide

scholarly journals Primary structure of a glycosylated DNA-binding domain in human plasma fibronectin.

1985 ◽  
Vol 260 (4) ◽  
pp. 2301-2306
Author(s):  
H Pande ◽  
J Calaycay ◽  
D Hawke ◽  
C M Ben-Avram ◽  
J E Shively
Keyword(s):  
Dna Binding ◽  
Human Plasma ◽  
Primary Structure ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Plasma Fibronectin

scholarly journals Functional mechanisms of MYRF DNA-binding domain mutations implicated in birth defects

2021 ◽  
Vol 296 ◽  
pp. 100612
Author(s):  
Chuandong Fan ◽  
Hongjoo An ◽  
Mohamed Sharif ◽  
Dongkyeong Kim ◽  
Yungki Park
Keyword(s):  
Dna Binding ◽  
Birth Defects ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Functional Mechanisms
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