PILOT_PROTEIN: identification of unmodified and modified proteins via high-resolution mass spectrometry and mixed-integer linear optimization.

A novel protein identification framework, PILOT_PROTEIN, has been developed to construct a comprehensive list of all unmodified proteins that are present in a living sample. It uses the peptide identification results from the PILOT_SEQUEL algorithm to initially determine all unmodified proteins within the sample. Using a rigorous biclustering approach that groups incorrect peptide sequences with other homologous sequences, the number of false positives reported is minimized.

Proteomic investigation of epigenetics in neuropsychiatric disorders: a missing link between genetics and behavior?

Neuropsychiatric disorders affect a large segment of the human population and result in large costs to society. The majority of such disorders have unknown underlying causes. Recent evidence suggests an important role for epigenetic regulation in the emergence of neuropsychiatric disease.

Quantitative proteomics reveals direct and indirect alterations in the histone code following methyltransferase knockdown.

Histones are highly conserved proteins that organize cellular DNA. These proteins, especially their N-terminal domains, are adorned with many post-translational modifications (PTMs) such as lysine methylation, which are associated with active or repressed transcriptional states. The lysine methyltransferase G9a and its interaction partner Glp1 can mono- or dimethylate histone H3 on lysine (H3K9me1 or me2); possible cross-talk between these modifications and other PTMs on the same or other histone molecules is currently uncharacterized.

Collective mass spectrometry approaches reveal broad and combinatorial modification of high mobility group protein A1a.

Transcriptional states are formed and maintained by the interaction and post-translational modification (PTM) of several chromatin proteins, such as histones and high mobility group (HMG) proteins. Among these, HMGA1a, a small heterochromatin-associated nuclear protein has been shown to be post-translationally modified, and some of these PTMs have been linked to apoptosis and cancer. In cancerous cells, HMGA1a PTMs differ between metastatic and nonmetastatic cells, suggesting the existence of an HMGA1a PTM code analogous to the "histone code.

Systems-wide proteomic characterization of combinatorial post-translational modification patterns.

Protein post-translational modifications (PTMs) have been widely shown to influence protein-protein interactions, direct subcellular location and transduce a variety of both internal and externally generated signals into cellular/phenotypic outcomes. Mass spectrometry has been a key tool for the elucidation of several types of PTMs in both qualitative and quantitative manners. As large datasets on the proteome-wide level are now being generated on a daily basis, the identification of combinatorial PTM patterns has become feasible.

A novel approach for untargeted post-translational modification identification using integer linear optimization and tandem mass spectrometry.

A novel algorithm, PILOT_PTM, has been developed for the untargeted identification of post-translational modifications (PTMs) on a template sequence. The algorithm consists of an analysis of an MS/MS spectrum via an integer linear optimization model to output a rank-ordered list of PTMs that best match the experimental data. Each MS/MS spectrum is analyzed by a preprocessing algorithm to reduce spectral noise and label potential complimentary, offset, isotope, and multiply charged peaks.

One-pot shotgun quantitative mass spectrometry characterization of histones.

Despite increasing applications of mass spectrometry (MS) to characterize post-translational modifications (PTMs) on histone proteins, most existing protocols are not properly suited to robustly measure them in a high-throughput quantitative manner. In this work, we expand on current protocols and describe improved methods for quantitative Bottom Up characterization of histones and their PTMs with comparable sensitivity but much higher throughput than standard MS approaches. This is accomplished by first bypassing off-line fractionation of histone proteins and working directly with total histones from a typical nuclei acid extraction.

High throughput characterization of combinatorial histone codes.

We present a novel method utilizing "saltless" pH gradient weak cation exchange-hydrophilic interaction liquid chromatography directly coupled to electron transfer dissociation (ETD) mass spectrometry for the automated on-line high throughput characterization of hypermodified combinatorial histone codes. This technique, performed on a low resolution mass spectrometer, displays an improvement over existing methods with an approximately 100-fold reduction in sample requirements and analysis time. The scheme presented is capable of identifying all of the major combinatorial histone codes present in a sample in a 2-h analysis.

Heterochromatin protein 1 is extensively decorated with histone code-like post-translational modifications.

Heterochromatin protein 1 (HP1) family members (alpha, beta, and gamma) bind histone H3 methylated at Lys-9, leading to gene silencing and heterochromatin formation. Several previous reports have suggested that HP1s are post-translationally modified, yet sites of modification have not yet been exhaustively determined. Here we perform the first comprehensive proteomic analysis of all HP1 isoforms using tandem mass spectrometry.