The FBI Laboratory

Smith, Wilder D.

doi:10.1021/ac0415511

Cited by 5 publications

(5 citation statements)

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“…After derivatization, peptide concentrations were determined using the bicinchoninic acid (BCA) assay 19. An angiotensin II standard curve was constructed and used to determine the total peptide concentration in the methylated and non‐methylated peptide samples.…”

Section: Methodsmentioning

confidence: 99%

An evaluation of the utility of in vacuo methylation for mass‐spectrometry‐based analyses of peptides

Laremore

Weber

Choma

2005

Rapid Comm Mass Spectrometry

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In vacuo trimethylation of the N-terminus of a lyophilized peptide with methyl iodide was previously reported to enhance the peptide's signal in matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) and to suppress alkali adduct formation in electrospray ionization mass spectrometry (ESI-MS). Both the signal enhancement and alkali adduct suppression observed for methylated peptides are believed to be due to the permanent positive charge on the N-terminus of the peptide resulting from the formation of a quaternary ammonium moiety. The present work evaluates the general utility of the in vacuo methylation procedure for the MS analysis of peptides, and specifically addresses the issue of whether the methylation of nucleophilic sites other than the N-terminal amine affects the MALDI signal of modified peptides. This work establishes that, although certain side-chain modifications are inevitable using present reaction conditions, the derivatization leads to significant MALDI-MS signal improvement. The experimental results demonstrate that the N-terminal trimethylammonium derivatives of peptides exhibit MALDI signals comparable to or exceeding those of arginine-containing standards such as angiotensin I. The advantages and limitations of the in vacuo methylation procedure are discussed.

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Section: Methodsmentioning

confidence: 99%

An evaluation of the utility of in vacuo methylation for mass‐spectrometry‐based analyses of peptides

Laremore

Weber

Choma

2005

Rapid Comm Mass Spectrometry

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“…An opaque band at the interface between the 5% and 30% layers was routinely harvested in fraction 4 and contained the lipid rafts. 125 I‐containing fractions were counted in a gamma counter, and aliquots were analyzed by the bicinchoninic acid method for total protein [31], and by immunoblotting for specific proteins resolved by electrophoresis on sodium dodecyl sulfate (SDS)–polyacrylamide gels.…”

Section: Methodsmentioning

confidence: 99%

Munc18‐2/syntaxin3 complexes are spatially separated from syntaxin3‐containing SNARE complexes

2003

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Exocytosis of mast cell granules requires a vesicularand plasma membrane-associated fusion machinery. We examined the distribution of SNARE membrane fusion and Munc18 accessory proteins in lipid rafts of RBL mast cells. SNAREs were found either excluded (syntaxin2), equally distributed between raft and non-raft fractions (syntaxin4, VAMP-8, VAMP-2), or selectively enriched in rafts (syntaxin3, SNAP-23). Syntaxin4-binding Munc18-3 was absent, whereas small amounts of the syntaxin3-interacting partner Munc18-2 consistently distributed into rafts. Cognate SNARE complexes of syntaxin3 with SNAP-23 and VAMP-8 were enriched in rafts, whereas Munc18-2/syntaxin3 complexes were excluded. This demonstrates a spatial separation between these two types of complexes and suggests that Munc18-2 acts in a step di¡erent from SNARE complex formation and fusion. ß

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“…The final pellet was resuspended in buffer B (0.25 M sucrose, 10 mM EDTA, 1 mM PMSF, 250 μM vitamin E, and 1 mM dithiothreitol, pH 7.5) and stored at −80°C for further analysis. The microsomal protein concentrations were determined using the bicinchoninic acid procedure [27].…”

Section: Methodsmentioning

confidence: 99%

Inflammatory prompts produce isoform‐specific changes in the expression of leukotriene B₄ ω‐hydroxylases in rat liver and kidney

Kalsotra¹,

Cui²,

Antonovic³

et al. 2003

FEBS Letters

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Cytochrome P450 (CYP) 4Fs metabolize leukotriene B 4 and other in£ammatory mediators in the arachidonic acid cascade. Here we show that lipopolysaccharide (LPS) treatment suppresses CYP4F4 and up-regulates CYP4F5 mRNA expression in rat liver whereas renal CYP4Fs are essentially unchanged. BaSO 4 treatment, in contrast, increases both hepatic and renal CYP4F expression levels. Thus, distinct regulatory mechanisms in CYP4F expression might operate under di¡erent in£ammatory prompts. To examine hepatic totipotency, primary hepatocytes were treated with varying doses of LPS resulting in decrease in all the CYP4F isoforms. Treatment of hepatocytes with 5 ng/ml of interleukin-1L L mimics the in vivo e¡ects of LPS on CYP4F expression. ß

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The FBI Laboratory

Cited by 5 publications

References 0 publications

An evaluation of the utility of in vacuo methylation for mass‐spectrometry‐based analyses of peptides

An evaluation of the utility of in vacuo methylation for mass‐spectrometry‐based analyses of peptides

Munc18‐2/syntaxin3 complexes are spatially separated from syntaxin3‐containing SNARE complexes

Inflammatory prompts produce isoform‐specific changes in the expression of leukotriene B₄ ω‐hydroxylases in rat liver and kidney

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The FBI Laboratory

Cited by 5 publications

References 0 publications

An evaluation of the utility of in vacuo methylation for mass‐spectrometry‐based analyses of peptides

An evaluation of the utility of in vacuo methylation for mass‐spectrometry‐based analyses of peptides

Munc18‐2/syntaxin3 complexes are spatially separated from syntaxin3‐containing SNARE complexes

Inflammatory prompts produce isoform‐specific changes in the expression of leukotriene B4 ω‐hydroxylases in rat liver and kidney

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Inflammatory prompts produce isoform‐specific changes in the expression of leukotriene B₄ ω‐hydroxylases in rat liver and kidney