title: Identification of ferredoxin II as a major calcium binding protein in the nitrogen-fixing symbiotic bacterium Mesorhizobium loti.
abstract: BACKGROUND
  Legumes establish with rhizobial bacteria a nitrogen-fixing symbiosis which is of the utmost importance for both plant nutrition and a sustainable agriculture. Calcium is known to act as a key intracellular messenger in the perception of symbiotic signals by both the host plant and the microbial partner. Regulation of intracellular free Ca(2+) concentration, which is a fundamental prerequisite for any Ca(2+)-based signalling system, is accomplished by complex mechanisms including Ca(2+) binding proteins acting as Ca(2+) buffers. In this work we investigated the occurrence of Ca(2+) binding proteins in Mesorhizobium loti, the specific symbiotic partner of the model legume Lotus japonicus.
  
  
  RESULTS
  A soluble, low molecular weight protein was found to share several biochemical features with the eukaryotic Ca(2+)-binding proteins calsequestrin and calreticulin, such as Stains-all blue staining on SDS-PAGE, an acidic isoelectric point and a Ca(2+)-dependent shift of electrophoretic mobility. The protein was purified to homogeneity by an ammonium sulfate precipitation procedure followed by anion-exchange chromatography on DEAE-Cellulose and electroendosmotic preparative electrophoresis. The Ca(2+) binding ability of the M. loti protein was demonstrated by (45)Ca(2+)-overlay assays. ESI-Q-TOF MS/MS analyses of the peptides generated after digestion with either trypsin or endoproteinase AspN identified the rhizobial protein as ferredoxin II and confirmed the presence of Ca(2+) adducts.
  
  
  CONCLUSIONS
  The present data indicate that ferredoxin II is a major Ca(2+) binding protein in M. loti that may participate in Ca(2+) homeostasis and suggest an evolutionarily ancient origin for protein-based Ca(2+) regulatory systems.
authors: Department of Biology, University of Padova, Via U. Bassi 58/B, 35131, Padova, Italy. roberto.moscatiello@unipd.it.
authors: Roberto
authors: R
authors: Moscatiello
authors: Department of Molecular Medicine, University of Padova, Viale G. Colombo 3, 35131, Padova, Italy. mattia.zaccarin@unipd.it.
authors: Mattia
authors: M
authors: Zaccarin
authors: Department of Biology, University of Padova, Via U. Bassi 58/B, 35131, Padova, Italy. flavia.ercolin@gmail.com.
authors: Flavia
authors: F
authors: Ercolin
authors: Department of Biomedical Sciences, University of Padova, Viale G. Colombo 3, 35131, Padova, Italy. ernesto.damiani@unipd.it.
authors: Ernesto
authors: E
authors: Damiani
authors: Department of Agronomy, Food, Natural Resources, Animals and Environment, DAFNAE, University of Padova, Viale dell'Università 16, 35020, Legnaro, Padova, Italy. squart@unipd.it.
authors: Andrea
authors: A
authors: Squartini
authors: Department of Molecular Medicine, University of Padova, Viale G. Colombo 3, 35131, Padova, Italy. antonella.roveri@unipd.it.
authors: Antonella
authors: A
authors: Roveri
authors: Department of Biology, University of Padova, Via U. Bassi 58/B, 35131, Padova, Italy. lorella.navazio@unipd.it.
authors: Lorella
authors: L
authors: Navazio
mesh_terms: D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D011232:Chemical Precipitation; D002852:Chromatography, Ion Exchange; D004586:Electrophoresis; D005288:Ferredoxins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D007526:Isoelectric Point; D059128:Mesorhizobium / Q000201:enzymology*; D009586:Nitrogen Fixation; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry
chemical_list: D002135:Calcium-Binding Proteins; D005288:Ferredoxins; D002118:Calcium
references: DNA Res. 2000 Dec 31;7(6):331-8
references: 11214968
references: Nat Rev Mol Cell Biol. 2000 Oct;1(1):11-21
references: 11413485
references: Trends Microbiol. 2002 Feb;10(2):87-93
references: 11827810
references: Brief Bioinform. 2002 Sep;3(3):265-74
references: 12230035
references: J Bacteriol. 2004 Apr;186(8):2439-48
references: 15060047
references: Mol Microbiol. 2004 Oct;54(2):291-7
references: 15469503
references: Proc Natl Acad Sci U S A. 2005 Apr 19;102(16):5744-8
references: 15811937
references: Proc Natl Acad Sci U S A. 2006 Jul 25;103(30):11334-9
references: 16849429
references: Cell Calcium. 2007 Oct-Nov;42(4-5):345-50
references: 17574670
references: Cell. 2007 Dec 14;131(6):1047-58
references: 18083096
references: DNA Res. 2008 Aug;15(4):227-39
references: 18511435
references: Biochem J. 1991 Aug 1;277 ( Pt 3):825-32
references: 1872815
references: J Bacteriol. 2009 Jun;191(12):4002-14
references: 19376875
references: Trends Microbiol. 2009 Oct;17(10):458-66
references: 19766492
references: BMC Microbiol. 2009 Sep 23;9:206
references: 19775463
references: J Bacteriol. 1991 Apr;173(7):2271-7
references: 2007551
references: J Biol Chem. 1991 Apr 15;266(11):7155-65
references: 2016321
references: Annu Rev Plant Biol. 2010;61:593-620
references: 20192754
references: New Phytol. 2010 Nov;188(3):814-23
references: 20738787
references: J Muscle Res Cell Motil. 1990 Dec;11(6):522-30
references: 2084148
references: J Biol Chem. 2011 Apr 8;286(14):12381-8
references: 21330362
references: Cold Spring Harb Perspect Biol. 2011 Mar 01;3(3):null
references: 21421925
references: Mol Biol Evol. 2012 Jan;29(1):91-100
references: 21680871
references: Annu Rev Genet. 2011;45:119-44
references: 21838550
references: Mol Plant Microbe Interact. 2012 Nov;25(11):1387-95
references: 22835276
references: Plant Physiol. 2012 Dec;160(4):2300-10
references: 23027664
references: Annu Rev Plant Biol. 2013;64:781-805
references: 23451778
references: Nat Rev Microbiol. 2013 Apr;11(4):252-63
references: 23493145
references: BMC Microbiol. 2013 Jul 31;13:180
references: 23898917
references: J Proteome Res. 2013 Nov 1;12(11):5323-30
references: 24041410
references: Planta. 1987 Aug;171(4):565-8
references: 24225721
references: Cell Calcium. 2015 Mar;57(3):151-65
references: 25555683
references: Biochem Biophys Res Commun. 1989 Dec 29;165(3):973-80
references: 2692569
references: Plants (Basel). 2013 Aug 26;2(3):530-40
references: 27137392
references: Biochemistry. 1987 Oct 6;26(20):6539-44
references: 3427023
references: Anal Biochem. 1973 Nov;56(1):43-51
references: 4128675
references: Anal Biochem. 1975 May 12;65(1-2):66-72
references: 48346
references: Nature. 1970 Aug 15;227(5259):680-5
references: 5432063
references: J Biol Chem. 1983 Sep 25;258(18):11267-73
references: 6193121
references: J Biochem. 1984 Feb;95(2):511-9
references: 6715311
references: Plant Physiol. 1995 Nov;109(3):983-90
references: 8552722
references: Electrophoresis. 1997 Mar-Apr;18(3-4):403-8
references: 9150918