Dataset Information

Human Adenovirus 5 (HAdV5), LC-MSMS

ABSTRACT: Using high-resolution mass spectrometry (MS)-based proteomics in combination with multiple protease digestion, we profiled, with on average 90% sequence coverage, all 13 viral proteins present in an human adenovirus (HAdV) vector. This in-depth profile provided multiple peptide-based evidence on intrinsic protease activity affecting several HAdV proteins. Next, the generated peptide library was used to develop a targeted proteomics method using selected reaction monitoring (SRM) aimed at quantitative profiling of the stoichiometry of all 13 proteins present in the HAdV. We also used this method to probe the release of specific virus proteins initiated by thermal stimulation, mimicking the early stage of HAdV disassembly during entry into host cells. We confirmed the copy numbers of the most well characterized viral capsid components and established the copy numbers for proteins whose stoichiometry has so far not been accurately defined. We also found that heating HAdV induces the complete release of the penton base and fiber proteins as well as a substantial release of protein VIII and VI. For these latter proteins, maturational proteolysis by the adenoviral protease (AVP) leads to the differential release of fragments with certain peptides being fully released and others largely retained in the AdV particles. This information is likely to be beneficial for the ongoing interpretation of high resolution cryoEM and X-ray electron density maps.

INSTRUMENT(S): LTQ Orbitrap, Q Exactive

ORGANISM(S): Human Adenovirus C Serotype 5 (hadv-5) (human Adenovirus 5)

SUBMITTER: Marco Benevento

LAB HEAD: Albert J.R. Heck

PROVIDER: PXD000591 | Pride | 2014-06-09

REPOSITORIES: Pride

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Publications

Adenovirus composition, proteolysis, and disassembly studied by in-depth qualitative and quantitative proteomics.

Benevento Marco M Di Palma Serena S Snijder Joost J Moyer Crystal L CL Reddy Vijay S VS Nemerow Glen R GR Heck Albert J R AJR

The Journal of biological chemistry 20140303 16

Using high-resolution MS-based proteomics in combination with multiple protease digestion, we profiled, with on average 90% sequence coverage, all 13 viral proteins present in an human adenovirus (HAdV) vector. This in-depth profile provided multiple peptide-based evidence on intrinsic protease activity affecting several HAdV proteins. Next, the generated peptide library was used to develop a targeted proteomics method using selected reaction monitoring (SRM) aimed at quantitative profiling of t ...[more]

PMID: 24591515

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Project description:The diagnosis of Kawasaki disease (KD) is often difficult to distinguish from adenovirus (HAdV) and Group A streptococcal disease (GAS). We sought to: 1) to define the KD transcriptional signature that can aid in the diagnosis of complete and incomplete KD in children; 2) to identify specific biomarkers that objectively discriminate between KD and other mimicking conditions, including HAdV and 3) to test the prognostic utility of GEP to determine response to IVIG therapy and development of coronary artery lesions (CAL). Methods: Blood RNA samples were analyzed from 76 pediatric patients with complete KD, 13 with incomplete KD, 19 patients with HAdV, 17 patients with GAS disease, and age- and sex-matched healthy controls (HC). We used class comparisons (MW p< 0.01, Benjamini-Hochberg, and 1.25 fold change filter), class prediction, modular analysis and MDTH analyses to define the specificity of the KD profiles and identify markers of severity. Results: Statistical group comparisons identified 7,899 genes differentially expressed in 39 complete KD patients versus HC (KD biosignature). This signature was validated in another 37 patients with complete KD and in 13 patients with incomplete KD. Modular analysis in children with complete KD demonstrated overexpression of inflammation, neutrophils, myeloid cell, coagulation cascade, and cell cycle genes. The KNN class prediction algorithm identified 25-classifier genes that differentiated children with KD vs HAdV infection in two independent cohorts of patients with 96% (95% CI [80%-99%]) sensitivity and 95% [74%-99%] specificity. MDTH scores in KD patients significantly correlated with the baseline c-reactive protein (R=0.29, p=0.008) and was four fold higher than in children with HAdV (p<0.01). In addition, KD patients that remained febrile 36 hours after treatment with IVIG (non-responders) demonstrated higher baseline, pre-treatment MDTH values compared with responders [12,290 vs. 5,572 respectively; p=0.009]. Conclusion: Transcriptional signatures can be used as a tool to discriminate between KD and HAdV infection, and may also provide prognostic information. 113 samples, no replicates, 9 HAdV, 57 cKD, 12 FUO, 13 GAS, 3 GAS/SF, 5 inKD, 14 healthy control

Dataset Information

Human Adenovirus 5 (HAdV5), LC-MSMS

Publications

Adenovirus composition, proteolysis, and disassembly studied by in-depth qualitative and quantitative proteomics.

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