Identification of low-abundance proteins in serum via the isolation of HSP72 complexes

Masako Tanaka, Masayuki Shiota, Takafumi Nakao, Ryo Uemura, Satoshi Nishi, Yasuyuki Ohkawa, Masaki Matsumoto, Maki Yamaguchi, Mayuko Osada-Oka, Azusa Inagaki, Katsuyuki Takahashi, Keiichi Nakayama, Min Gi, Yasukatsu Izumi, Katsuyuki Miura, Hiroshi Iwao

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

Heat shock protein 72 (HSP72) is an intracellular molecular chaperone that is overexpressed in tumor cells, and has also been detected in extracellular regions such as the blood. HSP72 forms complexes with peptides and proteins that are released from tumors. Accordingly, certain HSP72-binding proteins/peptides present in the blood of cancer patients may be derived from tumor cells. In this study, to effectively identify low-abundance proteins/peptides in the blood as tumor markers, we established a method for isolating HSP72-binding proteins/peptides from serum. Nine HSP72-specific monoclonal antibodies were conjugated to N-hydroxysulfosuccinimide-activated Sepharose beads (NHq) and used to isolate HSP72 complexes from serum samples. Precipitated proteins were then identified by LC-MS/MS analysis. Notably, this approach enabled the isolation of low-abundance proteins from serum without albumin removal. Moreover, by subjecting the serum samples of ten patients with multiple myeloma (MM) to NHq analysis, we identified 299 proteins present in MM HSP72 complexes, including 65 intracellular proteins. Among the intracellular proteins detected, 21 were present in all serum samples tested, while 11 were detected in both the conditioned media from cultured multiple myeloma cells and serum from MM patients. These results suggest that the NHq method can be applied to discover candidate tumor markers.

Original languageEnglish
Pages (from-to)214-221
Number of pages8
JournalJournal of Proteomics
Volume136
DOIs
Publication statusPublished - Mar 16 2016

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HSP72 Heat-Shock Proteins
Blood Proteins
Multiple Myeloma
Proteins
Tumors
Serum
Peptides
Blood
Tumor Biomarkers
Protein Binding
Neoplasms
Carrier Proteins
Cells
Myeloma Proteins
Molecular Chaperones
Conditioned Culture Medium
Serum Albumin
Sepharose
Albumins
Monoclonal Antibodies

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry

Cite this

Identification of low-abundance proteins in serum via the isolation of HSP72 complexes. / Tanaka, Masako; Shiota, Masayuki; Nakao, Takafumi; Uemura, Ryo; Nishi, Satoshi; Ohkawa, Yasuyuki; Matsumoto, Masaki; Yamaguchi, Maki; Osada-Oka, Mayuko; Inagaki, Azusa; Takahashi, Katsuyuki; Nakayama, Keiichi; Gi, Min; Izumi, Yasukatsu; Miura, Katsuyuki; Iwao, Hiroshi.

In: Journal of Proteomics, Vol. 136, 16.03.2016, p. 214-221.

Research output: Contribution to journalArticle

Tanaka, M, Shiota, M, Nakao, T, Uemura, R, Nishi, S, Ohkawa, Y, Matsumoto, M, Yamaguchi, M, Osada-Oka, M, Inagaki, A, Takahashi, K, Nakayama, K, Gi, M, Izumi, Y, Miura, K & Iwao, H 2016, 'Identification of low-abundance proteins in serum via the isolation of HSP72 complexes', Journal of Proteomics, vol. 136, pp. 214-221. https://doi.org/10.1016/j.jprot.2016.01.008
Tanaka, Masako ; Shiota, Masayuki ; Nakao, Takafumi ; Uemura, Ryo ; Nishi, Satoshi ; Ohkawa, Yasuyuki ; Matsumoto, Masaki ; Yamaguchi, Maki ; Osada-Oka, Mayuko ; Inagaki, Azusa ; Takahashi, Katsuyuki ; Nakayama, Keiichi ; Gi, Min ; Izumi, Yasukatsu ; Miura, Katsuyuki ; Iwao, Hiroshi. / Identification of low-abundance proteins in serum via the isolation of HSP72 complexes. In: Journal of Proteomics. 2016 ; Vol. 136. pp. 214-221.
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