title: Investigation of proteomic profiles of lamina of Ecklonia kurome (Laminariales): homology-based cross-species protein identification and analysis of the post-translational processing of vanadium-dependent bromoperoxidases using MALDI-TOF/TOF.
abstract: Proteomic profiles of the lamina of Ecklonia kurome Okamura, one of the Japanese dominant laminarialean kelps, were investigated by two-dimensional electrophoresis (2-DE) and MALDI-TOF/TOF. Due to the absence of E. kurome DNA or protein databases, homology-based cross-species protein identification was performed using a combination of three database-searching algorithms, Mascot peptide mass fingerprinting, Mascot MS/MS ion search, and mass spectrometry-based BLAST. Proteins were extracted from the lamina by an ethanol/phenol method and subjected to 2-DE (pI 4-7, 10 % polyacrylamide gel). More than 700 spots were detected in the 2-DE gel with CBB, and 93 spots (24 proteins) were successfully identified by MALDI-TOF/TOF and the cross-species database searching. The identified proteins mainly consisted of cytoplasmic carbohydrate metabolic enzymes, chloroplast proteins involved in photosynthesis, and haloperoxidases. Interestingly, vanadium-dependent bromoperoxidases (vBPO), which is thought to be involved in halogen uptake, synthesis of halogenated products, and detoxification of reactive oxygen species, were separated into at least 23 different spots. By comparing mass spectra, amino acid sequences predicted from tandem mass spectra and haloperoxidase activities of the vBPOs, we found that (1) at least two types of vBPOs were expressed in the lamina of E. kurome and (2) two pro-vBPOs might be activated by specific cleavage at N- and C-terminal regions.
authors.affiliation: Department of Genetic Engineering, Faculity of Biology-Oriented Science and Technology, Kinki University, Kinokawa, Wakayama 649-6493, Japan.
authors.forename: Kouhei
authors.initials: K
authors.lastname: Nagai
authors.forename: Koichi
authors.initials: K
authors.lastname: Morimoto
authors.forename: Haruka
authors.initials: H
authors.lastname: Ikegami
authors.forename: Hajime
authors.initials: H
authors.lastname: Kimura
authors.forename: Norishige
authors.initials: N
authors.lastname: Yotsukura
mesh_terms: D060365:Chloroplast Proteins / Q000235:genetics / Q000378:metabolism; D019295:Computational Biology / Q000379:methods; D015180:Electrophoresis, Gel, Two-Dimensional; D004798:Enzymes / Q000235:genetics / Q000378:metabolism*; D010449:Peptide Mapping; D010544:Peroxidases / Q000235:genetics / Q000378:metabolism; D000459:Phaeophyceae / Q000235:genetics / Q000378:metabolism*; D020543:Proteome / Q000235:genetics / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*; D013045:Species Specificity; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D053719:Tandem Mass Spectrometry
chemical_list: D060365:Chloroplast Proteins; D004798:Enzymes; D020543:Proteome; D010544:Peroxidases; C026195:bromide peroxidase
references.citation: Proteomics. 2010 Jun;10(11):2074-88
references.pmid: 20373519
references.citation: J Biol Chem. 1989 Nov 15;264(32):19287-92
references.pmid: 2553736
references.citation: Electrophoresis. 2003 Jul;24(14):2369-75
references.pmid: 12874872
references.citation: Plant Mol Biol. 2000 Jul;43(4):503-13
references.pmid: 11052202
references.citation: Science. 1973 Dec 7;182(4116):975-81
references.pmid: 17833778
references.citation: Mar Drugs. 2010 Mar 31;8(4):988-1010
references.pmid: 20479964
references.citation: J Proteome Res. 2010 May 7;9(5):2236-54
references.pmid: 20345176
references.citation: Proteomics. 2007 Jul;7(14):2318-29
references.pmid: 17623296
references.citation: Electrophoresis. 2008 Feb;29(3):672-81
references.pmid: 18228537
references.citation: J Biol Chem. 2003 Jun 27;278(26):23545-52
references.pmid: 12697758
references.citation: Proteomics. 2008 Jun;8(11):2266-84
references.pmid: 18452222
references.citation: J Phycol. 2008 Oct;44(5):1315-21
references.pmid: 27041728
references.citation: J Proteomics. 2008 Oct 7;71(4):461-72
references.pmid: 18656559
references.citation: J Biol Inorg Chem. 2008 Feb;13(2):257-69
references.pmid: 18008093
references.citation: Electrophoresis. 1999 Dec;20(18):3551-67
references.pmid: 10612281
references.citation: Proteomics. 2009 Feb;9(3):793-9
references.pmid: 19132686
references.citation: Proteomics. 2005 Jan;5(1):222-34
references.pmid: 15602773
references.citation: Proteomics. 2006 Mar;6(6):1897-907
references.pmid: 16479535
references.citation: Anal Chem. 2001 May 1;73(9):1917-26
references.pmid: 11354471
references.citation: Proc Natl Acad Sci U S A. 2008 May 13;105(19):6954-8
references.pmid: 18458346
references.citation: J Proteome Res. 2008 Feb;7(2):587-602
references.pmid: 18183946