Elucidating protein

Engineering zinc finger protein motifs for specific binding to double-stranded DNA is critical for targeted genome editing.

elucidating protein-46

Among many methods used to investigate protein/protein interactions, chemical cross-linking combined with mass spectrometry remains a vital experimental approach.

Mapping peptides modified by cross-linker provides clues about proteins' interacting domains.

It is postulated that the secretory isoform of gelsolin regulates several biological processes through interactions with proteins such as actin, fibronectin, vitamin D-binding protein, and unidentified receptors on the surface of eukaryotes; it also has been shown to self-assemble eventually leading to the formation of homo-multimers. We used four cross-linkers with arm length ranging from 7.7 to 21.7 Å and MALDI-TOF/TOF mass spectrometry as the analytical platform. TY - JOURT1 - Elucidating protein inter- and intramolecular interacting domains using chemical cross-linking and matrix-assisted laser desorption ionization-time of flight/time of flight mass spectrometry AU - Pottiez, Gwënaël AU - Ciborowski, Pawel PY - 2012/2/15Y1 - 2012/2/15N2 - Among many methods used to investigate protein/protein interactions, chemical cross-linking combined with mass spectrometry remains a vital experimental approach.

Results of this study show that MALDI-based mass spectrometry generates high quality data to show lysine residues modified by cross-linkers and combined with existing data based on crystallography (Protein Data Bank, PDB) can be used to discriminate between inter- and intramolecular linking. Mapping peptides modified by cross-linker provides clues about proteins' interacting domains.

One complication is that such modification may result from intra- but not intermolecular interactions.

Therefore, for overall data interpretation, a combination of results from various platforms is necessary.The rationale to study about zinc finger domain and its interaction with the DNA stems from the need to expatiate on the mechanisms by which the binding of transcription activators and repressors to the genome regulates the expression repertoire of all genes in a cell, hence focussing on its enormous scope in genome engineering.To exploit zinc finger proteins for genome manipulation, molecular and structural insights at the binding interface of zinc fingers and corresponding DNA targets are mandatory.Results of this study show that MALDI-based mass spectrometry generates high quality data to show lysine residues modified by cross-linkers and combined with existing data based on crystallography (Protein Data Bank, PDB) can be used to discriminate between inter- and intramolecular linking. The principles and most prominent recent applications of fluorescence correlation spectroscopy (FCS) are outlined, with special focus on intracellular studies, particularly protein dynamics and interactions in situ.Although there is no simple, general code for zinc finger protein–DNA recognition, selection strategies have been developed that allow these proteins to be designed to target almost any desired site on double-stranded DNA.

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