Phosphorylation and Acetylation of Acyl-CoA Synthetase- I.

Long chain acyl-CoA synthetase 1 (ACSL1) contributes 50 to 90% of total ACSL activity in liver, adipose tissue, and heart and appears to direct the use of long chain fatty acids for energy. Although the functional importance of ACSL1 is becoming clear, little is understood about its post-translational regulation. In order to investigate the post-translational modifications of ACSL1 under different physiological conditions, we overexpressed ACSL1 in hepatocytes, brown adipocytes, and 3T3-L1 differentiated adipocytes, treated these cells with different hormones, and analyzed the resulting phosphorylated and acetylated amino acids by mass spectrometry.

Acyl-coenzyme A synthetases in metabolic control.

Purpose Of Review: The 11 long-chain (ACSL) and very long chain acyl-coenzyme A (acyl-CoA) synthetases [(ACSVL)/fatty acid transport protein] are receiving considerable attention because it has become apparent that their individual functions are not redundant.

Recent Findings: Recent studies have focused on the structure of the acyl-CoA synthetases, their post-translational modification, their ability to activate fatty acids of varying chain lengths, and their role in directing fatty acids into different metabolic pathways. An unsettled controversy focuses on the ACSVL isoforms and whether these have both enzymatic and transport functions.

Investigations with O-linked protein glycosylations by matrix-assisted laser desorption/ionization Fourier transform ion cyclotron resonance mass spectrometry.

Posttranslational modifications such as glycosylation can play a fundamental role in signaling pathways that transform an ordinary cell into a malignant one. The development of a protocol to detect these changes in the preliminary stages of disease can lead to a sensitive and specific diagnostic for the early detection of malignancies such as ovarian cancer in which differential glycan patterns are linked to etiology and progression. Small variations in instrument parameters and sample preparation techniques are known to have significant influence on the outcome of an experiment.

Achieving augmented limits of detection for peptides with hydrophobic alkyl tags.

The wide range of protein concentrations found in biological matrixes presents a formidable analytical challenge in proteomics experiments. It is predicted that low-abundance proteins are the likely clinically relevant targets in disease-based proteomics analyses. To effectively analyze low-abundance proteins by electrospray ionization mass spectrometry, limits of detection must be improved upon.

Understanding the influence of post-excite radius and axial confinement on quantitative proteomic measurements using Fourier transform ion cyclotron resonance mass spectrometry.

Early studies of Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry (MS) explored many of the fundamental issues surrounding the potential of the technique to provide quantitative data. Improvements in instrument technology and the analysis of larger molecules in increasingly complex mixtures warrant not only a revisit to some of these earlier studies, but a more comprehensive examination of the influence of various instrument parameters on quantitative (absolute and relative) measurements in proteomics. We present a detailed examination of the role that acquisition time, excite voltage (i.

Accessible proteomics space and its implications for peak capacity for zero-, one- and two-dimensional separations coupled with FT-ICR and TOF mass spectrometry.

The number and wide dynamic range of components found in biological matrixes present several challenges for global proteomics. In this perspective, we will examine the potential of zero-dimensional (0D), one-dimensional (1D), and two-dimensional (2D) separations coupled with Fourier-transform ion cyclotron resonance (FT-ICR) and time-of-flight (TOF) mass spectrometry (MS) for the analysis of complex mixtures. We describe and further develop previous reports on the space occupied by peptides, to calculate the theoretical peak capacity available to each separations-mass spectrometry method examined.

Leveling response factors in the electrospray ionization process using a heated capillary interface.

Several investigators have observed a discrepancy in electrospray response of complementary strands from denatured DNA, which has been attributed to the difference in hydrophobicity between the two strands; the more hydrophobic species tend to have higher ion abundances. The implementation of a heated electrospray source has allowed us to "level" the electrospray response for two equimolar complementary strands with different hydrophobicities. As the temperature was increased, the ratio of ion abundances of the less hydrophobic noncoding strand to the more hydrophobic coding strand approached unity.