Separation and characterization of <i>Arabidopsis thaliana</i> proteins by two‐dimensional gel electrophoresis

Kamo, Masaharu; Kawakami, Takao; Miyatake, Norifumi; Tsugita, Akira

doi:10.1002/elps.1150160169

Cited by 91 publications

(56 citation statements)

References 19 publications

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“…The amino acid sequences of other two proteins could not be determined due to their blocked N termini. It has been well documented that many proteins have blocked N termini; consequently, Edman degradation does not proceed (e.g., Brown 1979;Tsugita et al 1994;Kamo et al 1995). The present results are not unusual.…”

Section: 48 ----------------------------------------------------supporting

confidence: 65%

Characterization of particulate proteins in Pacific surface waters

Saijo

Tanoue

2004

Limnology & Oceanography

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We investigated molecular characteristics of particulate proteins in Pacific surface waters using two-dimensional electrophoresis (2DE). Most proteinaceous materials estimated by dye-binding methods were characterized by the 2DE unresolved acidic materials with a broad range of molecular mass and the 2DE unresolved low molecular mass materials with a broad range of isoelectric point. The 2DE unresolved acidic and low molecular mass materials were considered to comprise peptides conjugated with acidic saccharides and degradation products (peptides) of proteins, respectively, which indicates that almost all proteins in living organisms failed to survive in detrital particulate organic matter (POM). Nevertheless, 23 discrete proteins were distinguished by the 2DE. Electrophoretic patterns of the discrete proteins indicated that they were a component of detrital POM. Three discrete proteins were subjected to N terminal amino acid sequence analysis. Two proteins out of three could not be determined because their N termini were blocked, and one protein was determined from the N terminus to the ninth amino acid residue. A homology search revealed that the N terminal amino acid sequence of the protein agreed completely with that of 70 kDa heat shock protein (HSP70) derived from photosynthetic organisms. HSP70 is a major member of the molecular chaperones that protect or repair proteins from damage under conditions of environmental stress. The occurrence of HSP70 in this study demonstrated that phytoplankton were able to induce the molecular chaperone(s). Clarification of factor(s) controlling induction of chaperones will enable us to assess the actual environmental stress on phytoplankton at the biomolecular level.The particulate organic matter (POM) in surface water is a complex mixture of living biomass and nonliving detritus. The contribution of living biomass to the bulk POM is generally 1 order of magnitude lower than that of detrital organic matter (e.g., Volkman and Tanoue 2002). The particulate combined amino acids (PCAA) are the largest identified fraction in surface POM (e.g., Wakeham et al. 1997), and dye-binding colorimetric methods demonstrated that proteinaceous materials were a major component of POM (Setchell 1981;Long and Azam 1996). Phytoplankton is a primary producer of organic matter in the sea, and most combined amino acids in phytoplankton are in the form of proteins. However, heterotrophic processing may convert cellular proteins of phytoplankton to other forms before its incorporation into detrital pool. At present the chemical nature of the detrital combined amino acids and proteinaceous materials is not well documented, and the processes by which organic matter produced by phytoplankton is transferred to detrital POM are not clear. The characterization of proteinaceous materials in POM at the molecular level is 1 Corresponding author (tanoue@ihas.nagoya-u.ac.jp). AcknowledgmentsWe thank the captains and crews of the RV Hakuho-maru KH93-4, KH99-3, RV Tansei-maru KT00-13, and TS Seisui-ma...

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Section: 48 ----------------------------------------------------supporting

confidence: 65%

Characterization of particulate proteins in Pacific surface waters

Saijo

Tanoue

2004

Limnology & Oceanography

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“…Proteins were extracted by the methods of Kamo et al (1995) 23) and Natera et al (2000), 15) with some minor modifications. To avoid biased sampling, whole roots and all nodules harvested were ground under liquid nitrogen, and then 1 g of the powder was suspended in cold acetone (À20 C) containing 10% w/v trichloroacetic acid and 0.07% w/v dithiothreitol (DTT).…”

Section: Methodsmentioning

confidence: 99%

Comparative Proteomic Analysis of Soybean Nodulation Using a Supernodulation Mutant, SS2-2

Lim

Park

Lee

et al. 2010

Bioscience, Biotechnology, and Biochemistry

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“…47 (2001) minal blockage than other species (Table 6). [44][45][46][47] Seventy-eight kDa glucose-regulated protein (shown in Table 1), which is identical to immunoglobulin heavy chain binding protein, is localized in the endoplasmic reticulum lumen 48) and belongs to the hsp70 family, sharing several family structural and biochemical characteristics.…”

Section: Analysis Of Phosphorylated Proteins Affected By Tcdd Exposurementioning

confidence: 99%

Proteome Analysis of the Effects of 2,3,7,8-Tetrachlorodibenzo-p-dioxin on Murine Testicular Leydig and Sertoli Cells.

Uchida

Ohashi

Morikawa

et al. 2001

JOURNAL OF HEALTH SCIENCE

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We treated testicular cell lines Leydig TM3 and Sertoli TM4 with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and used two-dimensional electrophoresis to investigate the resulting protein alterations. Cells were cultured in a medium containing 10 -5 to 10 nM TCDD for 4 hr, under which condition viability was not affected. Protein expression was compared semi-quantitatively by silver staining, by autoradiography of [35 S] methionine-labeled proteins, and by anti-phosphotyrosine antibody. In TM3, 34 protein spots were altered by TCDD, 26 of which were increased and 8 of which were decreased; in TM4, the amount of total protein appeared to be reduced and 19 protein spots were altered by TCDD, 12 of which were increased and 7 of which were decreased. Four of these altered proteins were identified by N-terminal protein microsequencing and by a homology search against protein databases. Whereas a pyruvate dehydrogenase E1 beta subunit was decreased by TCDD exposure, ATP synthase beta chain, mitochondrial matrix protein P1 and 78 kDa glucose-regulated protein were relatively increased. The precise role of these proteins in TCDD toxicity remains to be determined, but the observed alterations suggest the proteins to be important in the effects of TCDD on testicular cells.

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Separation and characterization of Arabidopsis thaliana proteins by two‐dimensional gel electrophoresis

Cited by 91 publications

References 19 publications

Characterization of particulate proteins in Pacific surface waters

Characterization of particulate proteins in Pacific surface waters

Comparative Proteomic Analysis of Soybean Nodulation Using a Supernodulation Mutant, SS2-2

Proteome Analysis of the Effects of 2,3,7,8-Tetrachlorodibenzo-p-dioxin on Murine Testicular Leydig and Sertoli Cells.

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