A Molecular Basis for the Presentation of Phosphorylated Peptides by HLA-B Antigens

Alpízar, Adán; Marino, Fabio; Ramos-Fernández, António; Lombardía, Manuel; Jekő, Anita; Pazos, Florencio; Paradela, Alberto; Santiago, César; Heck, Albert J. R.; Marcilla, Miguel

doi:10.1074/mcp.m116.063800

Cited by 56 publications

(76 citation statements)

References 54 publications

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“…Commonly, HLA class I and class II peptide isolation is achieved from cell lysates by IP. Cell suspensions or solid tissues are mechanically homogenised and lysed, employing non‐denaturing detergents, such as NP‐40, Triton X‐100, CHAPS, sodium deoxycholate or IGEPAL CA‐630 . Lysis buffers contain protease inhibitors to block degradation of HLA‐peptide complexes.…”

Section: Biochemical Isolation and Analysis Of Tumour Hla Ligandsmentioning

confidence: 99%

“…Lysis buffers contain protease inhibitors to block degradation of HLA‐peptide complexes. Optionally, phosphatase inhibitors can be added to prevent dephosphorylation of phosphopeptides . In some cases, iodacetamide is added as well to alkylate cysteins, thereby inhibiting disulphide bond formation.…”

Section: Biochemical Isolation and Analysis Of Tumour Hla Ligandsmentioning

confidence: 99%

“…Elution of HLA complexes can be achieved either through treatment with a strong acid, such as 0·1–0·2% TFA, 10% acetic acid or with 0·1–0·2 N acetic acid followed by heat denaturation . Peptides are frequently separated from HLA fragments through ultrafiltration, employing a molecular weight filter.…”

Section: Biochemical Isolation and Analysis Of Tumour Hla Ligandsmentioning

confidence: 99%

“…Therefore, enrichment prior to MS increases identification rates. Enrichment strategies are frequently variations of the immobilised metal affinity chromatography (IMAC), in which metal ions such as Ti 4+ or Fe 3+ chelate phosphonate groups . Derivatisation of the carboxy‐termini to methyl esters has been employed to decrease unspecific interaction of acidic peptides with metal ions by some groups .…”

Section: Biochemical Isolation and Analysis Of Tumour Hla Ligandsmentioning

confidence: 99%

“…Derivatisation of the carboxy‐termini to methyl esters has been employed to decrease unspecific interaction of acidic peptides with metal ions by some groups . Metal oxide affinity chromatography (MOAC) follows a similar enrichment mechanism, but uses metal oxides such as Ti 2 O as solid phase . SCX chromatography is often used as a primary or secondary prefractionation strategy in combination with either IMAC or MOAC enrichment …”

Section: Biochemical Isolation and Analysis Of Tumour Hla Ligandsmentioning

confidence: 99%

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Mapping the tumour human leukocyte antigen (HLA) ligandome by mass spectrometry

2018

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The entirety of human leukocyte antigen (HLA)-presented peptides is referred to as the HLA ligandome of a cell or tissue, in tumours often termed immunopeptidome. Mapping the tumour immunopeptidome by mass spectrometry (MS) comprehensively views the pathophysiologically relevant antigenic signature of human malignancies. MS is an unbiased approach stringently filtering the candidates to be tested as opposed to epitope prediction algorithms. In the setting of peptide-specific immunotherapies, MS-based strategies significantly diminish the risk of lacking clinical benefit, as they yield highly enriched amounts of truly presented peptides. Early immunopeptidomic efforts were severely limited by technical sensitivity and manual spectra interpretation. The technological progress with development of orbitrap mass analysers and enhanced chromatographic performance led to vast improvements in mass accuracy, sensitivity, resolution, and speed. Concomitantly, bioinformatic tools were developed to process MS data, integrate sequencing results, and deconvolute multi-allelic datasets. This enabled the immense advancement of tumour immunopeptidomics. Studying the HLA-presented peptide repertoire bears high potential for both answering basic scientific questions and translational application. Mapping the tumour HLA ligandome has started to significantly contribute to target identification for the design of peptide-specific cancer immunotherapies in clinical trials and compassionate need treatments. In contrast to prediction algorithms, rare HLA allotypes and HLA class II can be adequately addressed when choosing MS-guided target identification platforms. Herein, we review the identification of tumour HLA ligands focusing on sources, methods, bioinformatic data analysis, translational application, and provide an outlook on future developments.

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Section: Biochemical Isolation and Analysis Of Tumour Hla Ligandsmentioning

confidence: 99%

Section: Biochemical Isolation and Analysis Of Tumour Hla Ligandsmentioning

confidence: 99%

Section: Biochemical Isolation and Analysis Of Tumour Hla Ligandsmentioning

confidence: 99%

Section: Biochemical Isolation and Analysis Of Tumour Hla Ligandsmentioning

confidence: 99%

Section: Biochemical Isolation and Analysis Of Tumour Hla Ligandsmentioning

confidence: 99%

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Mapping the tumour human leukocyte antigen (HLA) ligandome by mass spectrometry

2018

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Gaining Insight Into Posttranslationally Modified HIV Antigens and Their Underlying Characteristics

Marino

2018

Proteomics

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Mass spectrometry (MS)-based immunopeptidomics has developed as one of the leading methodologies for comprehensive characterization of in vivo presented human leukocyte antigen (HLA)-bound peptides. Unveiling the identity of HLA-bound peptides derived from diseased cells is crucial to gain knowledge on the constitution of efficient disease-specific T cell responses. The HLA-presented peptidome reflects the status of the cellular proteome, hence disease-related aberrations of posttranslational modifications (PTMs) might lead to presentation of peptides harboring PTMs. Therefore, characterization of HLA-bound PTM peptides could shed light on their relevance in immune and disease processes. In this issue, Ramarathinam et al. investigate the presentation of HIV envelope (HIVenv) peptides bound to the HLA-B*57:01 allele. Among these peptides, the authors specifically focused on a kynurenine-modified peptide. To this end, they characterize the possible origin of the kynurenine modification, its effect on HLA binding affinity, stability, conformation within the complex, and its immunogenicity compared to the native counterpart.

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Immunopeptidomics Special Issue

Ternette

Purcell

2018

Proteomics

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The use of peptides to infer protein expression is very familiar to all of those who undertake bottom-up proteomics studies of cells, tissues, and fluids. Remarkably, an analogous process is used by the immune system to report on the presence of infectious microorganisms, malignancy, and other abnormalities within the body. This process known as antigen processing and presentation relies on the selection and presentation of peptides on the surface of cells as a bound complex with major histocompatibility complex (MHC) molecules. The isolation and characterization of these MHC-bound peptides, predominantly through the use of mass spectrometry, has been coined immunopeptidomics and is the subject of this special issue.Immunopeptidomics as a field has rapidly matured in the past decade and has moved from being the domain of just a handful of specialized laboratories to now being more generally applied by the proteomics and immunology communities. The development of immunoproteomics was pioneered by the groups of Don Hunt with Victor Englehard and Alessandro Sette [1][2][3][4][5] and Hans-Georg Rammensee. [6][7][8][9] Over the years, the protocols developed by these groups have been refined but the basic premise of isolating peptide-MHC complexes under near-native conditions and the analysis of the bound peptide cargo primarily by LC-MS/MS remain unchanged. What has changed is the instrumentation and sensitivity of the measurements which have allowed analysis of thousands of peptides using this technique. With such information-rich and deep datasets reporting on peptide epitope display by a variety of different cell types and tissues have come profound insights into fundamental mechanisms of antigen presentation (e.g., refs. 2,4,10-17) and how this may be perturbed by various environmental or infectious agents (e.g., refs. 18-20). The maturation of immunopeptidomics has also facilitated the identification of T cell epitopes in a variety of diseases including autoimmunity (e.g., refs. 21-24), cancer (e.g., refs. 25-31), and infectious disease (e.g., refs. 32-35). It has revealed the critical role that post-translational modification (e.g., refs. 24,30,31,36-40) has in the immune response and allowed enumeration of specific epitopes with great accuracy and precision (e.g., refs. 41-43).In this special issue, we have collected contributions that apply immunopeptidomics in the context of infectious disease, [44,45] cancer, [46] and graft versus host disease. [47] Furthermore, the development of HLA binding prediction and the use of highquality mass spectrometric data are discussed in two review articles. [48,49] Enrichment strategies prior to MS analysis are compared for suitability of antigen discovery [50] and alternative instrument acquisition strategies to the standard data-dependent acquisition (MS3, [45] PRM, [47] EThcD [51] , and DIA [52] ) are discussed. An important outcome of a recent meeting that focused on immunopeptidomics and the establishment of the Human Immunopeptidome project [53] is a series of guide...

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A Molecular Basis for the Presentation of Phosphorylated Peptides by HLA-B Antigens

Cited by 56 publications

References 54 publications

Mapping the tumour human leukocyte antigen (HLA) ligandome by mass spectrometry

Mapping the tumour human leukocyte antigen (HLA) ligandome by mass spectrometry

Gaining Insight Into Posttranslationally Modified HIV Antigens and Their Underlying Characteristics

Immunopeptidomics Special Issue

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