Probing Rhodopsin−Transducin Interactions by Surface Modification and Mass Spectrometry

Wang, Xin; Kim, Sung Hoon; Ablonczy, Zsolt; Crouch, Rosalie K.; Knapp, Daniel R.

doi:10.1021/bi049642f

Cited by 42 publications

(49 citation statements)

References 39 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Numerous MS-based covalent labeling experiments have been conducted on membrane proteins [28,29], e.g., for topologic investigations [30], as well as for monitoring conformational changes [29,[31][32][33][34] and binding interactions [35][36][37].…”

mentioning

confidence: 99%

Site-directed mutagenesis combined with oxidative methionine labeling for probing structural transitions of a membrane protein by mass spectrometry

Pan

Brown

Konermann

2010

J. Am. Soc. Mass Spectrom.

View full text Add to dashboard Cite

mentioning

confidence: 99%

Site-directed mutagenesis combined with oxidative methionine labeling for probing structural transitions of a membrane protein by mass spectrometry

Pan

Brown

Konermann

2010

J. Am. Soc. Mass Spectrom.

View full text Add to dashboard Cite

“…Others have also successfully used acetic anhydride for structure determinations [9,10]. Recently, several new reagents and labels have emerged that target solvent accessible lysines: N-hydroxysuccinimido-biotin (NHS-biotin) [11,12], sulfosuccinimidyl acetate [13] and S-methylthioacetimidate [14]. Especially, the use of NHS-ester chemistry in lysine labeling has proven to be efficient and reliable.…”

mentioning

confidence: 99%

Analysis of protein-protein interaction surfaces using a combination of efficient lysine acetylation and nanoLC-MALDI-MS/MS applied to the E9:Im9 bacteriotoxin—immunity protein complex

Scholten

Visser

Heuvel

et al. 2006

J. Am. Soc. Mass Spectrom.

View full text Add to dashboard Cite

To understand how proteins perform their function, knowledge about their structure and dynamics is essential. Here we use a combination of an efficient chemical lysine acetylation reaction and nanoLC-MALDI tandem mass spectrometry to probe the accessibility of every lysine residue in a protein complex. To demonstrate the applicability of this approach, we studied the interaction between the DNase domain of Colicin E9 (E9) and its immunity protein Im9. Free E9 and E9 in complex with Im9 were rapidly acetylated, followed by proteolytic digestion and analysis by LC-MALDI-TOF/TOF MS/MS. Acetylated peptides could be filtered out of the complex peptide mixtures using selective ion chromatograms of the specific immonium marker ions. Additionally, isobaric acetylated peptides, acetylated at different sites, could be separated by their LC retention times. The combination of LC and MALDI-TOF/TOF MS/MS provided information about the amount of acetylation on each individual lysine even for peptides containing several lysine residues. In general, our data agree well with those derived from the crystal structure of E9 and the E9:Im9 complex. Interestingly, next to in the binding interface expected lysines, K89 and K97, two from the crystal structure data unexpected lysines, K81 and K76, were observed to become less exposed upon Im9 binding. Moreover, K55 and K63, positioned in the predicted DNA binding region, were also found to be less accessible upon Im9 binding. These findings may illustrate some of the described differences in the solution-phase structure of the E9:Im9 complex compared with the crystal structure. (J Am Soc Mass Spectrom 2006, 17, 983-994)

show abstract

“…SNA was chosen previously to probe the surface Lys residue on the plasma transglutaminase, Factor XIIIA 20 and several other enzymes. 21 Another pharmacologic acetylating agent, aspirin (acetyl salicylic; ASA), which is a common anti-inflammatory agent, acetylated Lys residues in several molecules including antithrombin III, 22 hemoglobin, 23,24 fibrinogen 25 and BSA. 26 Since ASA can be administered orally to humans, it could serve as a practical method to modulate the activity of TGM2.…”

Section: Resultsmentioning

confidence: 99%

Human tissue transglutaminase is inhibited by pharmacologic and chemical acetylation

2010

View full text Add to dashboard Cite

Human tissue transglutaminase (TGM2) is implicated in the pathogenesis of several neurodegenerative disorders including Alzheimer's, Parkinson's and expanded polyglutamine (polyQ) diseases. TGM2 promotes formation of soluble and insoluble high molecular weight aggregates by catalyzing a covalent linkage between peptide-bound Q residues in polyQ proteins and a peptide-bound Lys residue. Therapeutic approaches to modulate the activity of TGM2 are needed to proceed with studies to test the efficacy of TGM2 inhibition in disease processes. We investigated whether acetylation of Lys-residues by sulfosuccinimidyl acetate (SNA) or aspirin (ASA) would alter the crosslinking activity of TGM2. Acetylation by either SNA and/or ASA resulted in a loss of >90% of crosslinking activity. The Lys residues that were critical for inhibition were identified by mass spectrometry as Lys . Hence, acetylation of Lys-residues may modulate the enzymatic function of TGM2 in vivo and offer a novel approach to treatment of TGM2 mediated disorders.

show abstract

Probing Rhodopsin−Transducin Interactions by Surface Modification and Mass Spectrometry

Cited by 42 publications

References 39 publications

Site-directed mutagenesis combined with oxidative methionine labeling for probing structural transitions of a membrane protein by mass spectrometry

Site-directed mutagenesis combined with oxidative methionine labeling for probing structural transitions of a membrane protein by mass spectrometry

Analysis of protein-protein interaction surfaces using a combination of efficient lysine acetylation and nanoLC-MALDI-MS/MS applied to the E9:Im9 bacteriotoxin—immunity protein complex

Human tissue transglutaminase is inhibited by pharmacologic and chemical acetylation

Contact Info

Product

Resources

About