Large-Scale Examination of Factors Influencing Phosphopeptide Neutral Loss during Collision Induced Dissociation.

Collision-induced dissociation (CID) remains the predominant mass spectrometry-based method for identifying phosphorylation sites in complex mixtures. Unfortunately, the gas-phase reactivity of phosphoester bonds results in MS/MS spectra dominated by phosphoric acid (H3PO4) neutral loss events, suppressing informative peptide backbone cleavages. To understand the major drivers of H3PO4 neutral loss, we performed robust nonparametric statistical analysis of local and distal sequence effects on the magnitude and variability of neutral loss, using a collection of over 35,000 unique phosphopeptide MS/MS spectra. In contrast to peptide amide dissociation pathways, which are strongly influenced by adjacent amino acid side chains, we find that neutral loss of H3PO4 is affected by both proximal and distal sites, most notably basic residues and the peptide N-terminal primary amine. Previous studies have suggested that protonated basic residues catalyze neutral loss through direct interactions with the phosphate. In contrast, we find that nearby basic groups decrease neutral loss regardless of mobility class, an effect only seen by stratifying spectra by charge-mobility. The most inhibitory bases are those immediately N-terminal to the phosphate, presumably because of steric hindrances in catalyzing neutral loss. Further evidence of steric effects is shown by the presence of proline, which can dramatically reduce the presence of neutral loss when between the phosphate and a possible charge donor. In mobile proton spectra, the N-terminus is the strongest predictor of high neutral loss, with proximity to the N-terminus essential for peptides to exhibit the highest levels of neutral loss.

Development and process evaluation of a primary care internet-based intervention to prevent depression in emerging adults.

BACKGROUND: Primary care is a potential setting for implementation of depression prevention interventions using cognitive behavioral therapy (CBT) and interpersonal psychotherapy (IPT). The purpose of this study was to develop and conduct a process evaluation of a primary care/ Internet-based intervention that addresses key dissemination barriers in a community setting. METHOD: We used an interdisciplinary team of investigators in a multistep intervention development process among a sample of primary care patients (aged 18 to 24 years). The intervention included an initial primary care motivational interview to engage the participant, 11 Internet-based modules based on CBT (to counter pessimistic thinking) and IPT (to activate social networks and strengthen relationship skills), and a follow-up motivational interview in primary care to enhance behavior change. Each component of the intervention was rated with regard to dissemination barriers of (1) fidelity, (2) motivation, (3) dose, (4) perceived helpfulness (rated on a Likert scale), and (5) potential costs. The study was conducted from April through June of 2004. RESULTS: Fidelity checklist and serial reviews were satisfactory (100% core concepts translated into intervention). Key motivations for participation included (1) risk reduction, (2) intervention effectiveness, (3) "resiliency," and (4) altruism. In terms of dose, 13 of 14 participants engaged the Internet-based components, completing a mean of 7.2 modules (SD = 3.9). The 2 primary care interviews and the self-assessment and resiliency modules received the highest helpfulness ratings. The duration of the 2 motivational interviews was approximately 17-18 minutes, which is similar to a typical primary care visit. CONCLUSIONS: By using multidisciplinary teams and incorporating the opinions of potential users, complex preventive mental health interventions can be translated into primary care settings with adequate fidelity, motivation, dose, and perceived helpfulness, and at a reasonably low cost.