Da a A icle
Da a se o he p o eomic in en o y and
quan i a i e analysis o chicken eggshell ma ix
p o eins du ing he p ima y e en s o eggshell
mine aliza ion and he ac i e g ow h phase
o calcifica ion
Pauline Ma ie
a
, Valé ie Labas
b
, Au élien B ionne
a
,
G égoi e Ha ichaux
b
, Ch is elle Henneque -An ie
a
, Alejand o
B. Rod iguez-Na a o
c
, Y es Nys
a
, Joël Gau on
a,
n
a
INRA, UR83 Reche ches a icoles, Fonc ion e Régula ion des p o éines de l'œu , F-37380 Nouzilly, F ance
b
INRA, UMR INRA85, UMR CNRS 7247; Uni e si é de Tou s; IFCE, Physiologie de la Rep oduc ion e des
Compo emen s, Pla e- o me d'Analyse In ég a i e des Biomolécules, Labo a oi e de Spec omé ie de Masse,
F-37380 Nouzilly, F ance
c
Depa men o de Mine alogia y Pe ologia, Uni e sidad de G anada, Campus Fuen e Nue a s/n, 18002
G anada, Spain
a icle in o
A icle his o y:
Recei ed 15 June 2015
Recei ed in e ised o m
26 June 2015
Accep ed 29 June 2015
A ailable online 7 July 2015
Keywo ds:
Chicken
Eggshell
Biomine alisa ion
Quan i a i e p o eomics
Eggshell ma ix
abs ac
Chicken eggshell is a biomine al composed o 95% calci e calcium
ca bona e mine al and o 3.5% o ganic ma ix p o eins. The
assembly o mine al and i s s uc u al o ganiza ion is con olled
by i s o ganic ma ix. In a ecen s udy [1], we ha e used
quan i a i e p o eomic, bioin o ma ic and unc ional analyses o
explo e he dis ibu ion o 216 eggshell ma ix p o eins a ou key
s ages o shell mine aliza ion defined as: (1) widesp ead deposi-
ion o amo phous calcium ca bona e (ACC), (2) ACC ans o ma-
ion in o c ys alline calci e agg ega es, (3) o ma ion o la ge
calci e c ys al uni s and (4) apid g ow h o calci e as columna
s uc u e wi h p e e en ial c ys al o ien a ion. The cu en a icle
de ailed he quan i a i e analysis pe o med a he ou s ages o
Con en s lis s a ailable a ScienceDi ec
jou nal homepage: www.else ie .com/loca e/dib
Da a in B ie
h p://dx.doi.o g/10.1016/j.dib.2015.06.019
2352-3409/&2015 The Au ho s. Published by Else ie Inc. This is an open access a icle unde he CC BY license
(h p://c ea i ecommons.o g/licenses/by/4.0/).
DOI o o iginal a icle: h p://dx.doi.o g/10.1016/j.jp o .2015.05.034
n
Co esponding au ho .
E-mail add ess: [email p o ec ed] (J. Gau on).
Da a in B ie 4 (2015) 430–436
shell mine aliza ion o de e mine he p o eins which a e he mos
abundan . Addi ionally, we epo ed he en iched GO e ms and
desc ibed he p esence o 35 an imic obial p o eins equally
dis ibu ed a all s ages o keep he egg ee o bac e ia and o
81 p o eins, he unc ion o which could no be asc ibed.
&2015 The Au ho s. Published by Else ie Inc. This is an open
access a icle unde he CC BY license
(h p://c ea i ecommons.o g/licenses/by/4.0/).
Specifica ions able
Subjec a ea Biology
Mo e specific
subjec a ea
Lis and pu a i e unc ions o eggshell ma ix p o eins p esen a ini ial and mid phases o eggshell
o ma ion.
Type o da a Raw and p ocessed/analyzed mass spec ome y da a ob ained by nanoliquid ch oma og aphy
combined o high esolu ion andem mass spec ome y,.xls ables wi h iden ified/ alida ed and
quan ified p o eins ables wi h in eg a i e and unc ional analysis o p o ein sequences.
How da a was
acqui ed
LC–MS/MS using a LTQ O bi ap Velos mass spec ome e .
Da a o ma Raw da a: aw.mzml and P ocessed and analyzed da a using Masco Sea ch engine:.da .
Analyzed: Fu he assembled sequences using Clus al Omega mul i-alignmen algo i hm and
BLASTþsui e. GO e ms ex ac ed om sequences and en ichmen de e mina ion.
Expe imen al ac o s S age o eggshell o ma ion.
Expe imen al
ea u es
Eggshell ma ix samples we e collec ed a ou s ages o eggshell mine aliza ion and diges ed in gel
using ypsin. Resul ing pep ides we e analyzed by LC–MS/MS and u he ea ed using da a mining
and bioin o ma ic analysis.
Da a sou ce loca ion Nouzilly, F ance, INRA Cen e Val de Loi e.
Da a accessibili y Da a ha e been deposi ed in o he P o eomeXchange Conso ium [2] ia he PRIDE pa ne
eposi o y wi h he da ase iden ifie PXD001450.
Value o he da a [desc ibe in 3–5 bulle ed poin s why his da a is o alue o he scien ific
communi y]

P o eomic analysis o 216 chicken eggshell ma ix p o eins.

Gene On ology e ms en ichmen s in es iga ing po en ial unc ions o eggshell ma ix p o eins.

Quan i a i e da a on p o ein abundances acco ding o s ages o eggshell o ma ion.

Anno a ion on quan ified eggshell ma ix p o eins.
1. Collec ion o eggshell ma ix samples and p epa a ion o MS analyses
Eggs we e collec ed on b own-laying hens a 5, 6, 7 and 16 h a e p e ious o iposi ion as
desc ibed in [1]. These ime in e als co espond espec i ely o he p ima y e en s o mine aliza ion
wi h widesp ead deposi ion o amo phous calcium ca bona e (ACC), ACC ans o ma ion in o
c ys alline calci e agg ega es, o ma ion o la ge calci e c ys al uni s and apid g ow h o calci e and
de elopmen o a columna s uc u e wi h p e e en ial c ys al o ien a ion [3]. Eggs we e b oken and
o ming eggshells we e washed wi h wa e , ai d ied and s o ed a 20 1C un il p o ein ex ac ion.
Eggshell ma ix p o eins we e ex ac ed as desc ibed in [1].
A o al o 24 indi idual eggshell p o ein ex ac s we e used. Six samples collec ed a he same ime
poin we e pooled in equal amoun s o each ime (5 h p.o., 6 h p.o., 7 h p.o. and 16 h p.o.). The ou
pooled samples (51
m
g o p o eins/sample) we e ac iona ed on a 4–20% SDS-PAGE gel
(8.3 cm 7.3 cm 1.5 mm). P o eins we e s ained wi h Coomassie blue and he en i e SDS-PAGE
lanes we e sec ioned in o 15 bands o each indi idual pooled sample. Excised p o eins we e in-gel
P. Ma ie e al. / Da a in B ie 4 (2015) 430–436 431
diges ed wi h bo ine ypsin (Roche Diagnos ics GmbH, Mannhiem, Ge many) as p e iously desc ibed
[4] and analyzed by nanoscale liquid ch oma og aphy- andem mass spec ome y (nanoLC–MS/MS).
2. Nano LC MS/MS analyses
All expe imen s we e pe o med on a linea ion ap Fou ie T ans o m Mass Spec ome e (FT-MS)
LTQ O bi ap Velos (The mo Fishe Scien ific, B emen, Ge many) coupled o an Ul ima e
s
3000 RSLC
Ul a-High P essu e Liquid Ch oma og aphe (Dionex, Ams e dam, The Ne he lands) as p e iously
desc ibed in [1,4]. Raw da a files we e con e ed o MGF as p e iously desc ibed [4]. The iden ifica ion
o p o eins was es ablished using MASCOT sea ch engine ( 2.3, Ma ix Science, London, UK). The
pep ide and agmen masses ob ained we e ma ched au oma ically agains he cho da a sec ion o n
NCBI da abase (2132453 sequences, downloaded on 2013/05/13). Enzyme specifici y was se o
ypsin wi h wo missed clea ages using ca bamidome hylcys eine, oxida ion o me hionine and N-
e minal p o ein ace yla ion as a iable modifica ions. The ole ance o he ions was se o 5 ppm o
pa en and 0.8 Da o agmen ion ma ches. Masco esul s ob ained om he a ge and decoy
da abase we e inco po a ed in Sca old 3 so wa e ( e sion 3.4, P o eome So wa e, Po land, USA).
Pep ide iden ifica ions we e accep ed i hey could be es ablished a g ea e han 95.0% p obabili y as
specified by he Pep ide P ophe algo i hm [5]. Pep ides we e conside ed dis inc i hey di e ed in
sequence. P o ein iden ifica ions we e accep ed i hey could be es ablished a g ea e han 95.0%
p obabili y as specified by he P o ein P ophe algo i hm [6] and con ained a leas one sequence-
unique pep ide o global in en o y and a leas wo sequence-unique pep ides o quan i a i e
analysis. A alse disco e y a e was calcula ed as o1% a he pep ide o p o ein le el. The abundance
o iden ified p o eins was es ima ed by calcula ing he emPAI using Sca old 4 Qþso wa e ( e sion
4.2, P o eome So wa e, Po land, USA). Addi ionally, label- ee quan i a i e p o eomic analyses based
on spec al coun ing me hod, Sca old 3 Qþso wa e ( e sion 3.4, P o eome So wa e, Po land, USA)
was used o quan i y he p o eins a he ou di e en s ages o calcifica ion. All p o eins wi h g ea e
han wo sequence-unique pep ides, iden ified in da abase wi h high confidence we e conside ed o
p o ein quan ifica ion. To elimina e quan i a i e ambigui y wi hin p o ein g oups, we igno ed all he
spec a ma ching any pep ide which is sha ed be ween p o eins. The eby, quan ifica ion pe o med
wi h no malized spec al coun s was ca ied ou on dis inc p o eins.
The mass spec ome y p o eomic da a ha e been deposi ed in o he P o eomeXchange
Conso ium [2] ia he PRIDE pa ne eposi o y wi h he da ase iden ifie PXD001450.
3. Da a se analysis o a ian eggshell ma ix p o eins
A o al o 261 p o eins we e iden ified using n NCBI da abase. Ke a ins and bo ine ypsin we e
elimina ed om he lis as hey appea ed o be con aminan s o esul ing om he diges ion p ocess.
P o ein sequences we e aligned o elimina e all edundancies. P o ein g oups we e de e mined using
Clus al Omega mul i-alignmen algo i hm [7]. Sequences we e blas ed agains n NCBI da abase
limi ed o Gallus gallus axon using he blas p p og am (BLASTþsui e) [8]. This was pe o med using R
language (h p://c an. -p ojec .o g).
A o al o 216 non- edundan eggshell ma ix p o ein sequences we e iden ified. The esul ing file
(Supplemen a y Table 1) is made o En ezGene Ids, GI numbe s, p o ein symbols, sho desc ip ions,
in o ma ion on hei p e ious iden ifica ion in he shell and in he u e ine fluid, mean quan i a i e
alues and emPAI alues a each ime poin . We compa ed ou lis wi h he p e iously published
p o eomes o eggshell ma ix p o eins [9–15]. Ou o he 216 p o eins, 24 p o eins a e no el compa ed
o p e ious s udies and a e highligh ed (Supplemen a y Table 1).
In o de o de e mine he po en ial unc ions o he 216 eggshell ma ix p o eins, 3231 GO e ms
we e ex ac ed om he p o ein sequences using GORe ie e (h p://agbase.mss a e.edu/).
Ca eGO ize (h p://www.animalgenome.o g/ ools/ca ego/) was hen used o g oup he GO e ms in
91 di e en pa en e m ca ego ies using a GO slim2 me hod. Finally, GO e ms en ichmen s we e
de e mined using Gene se en ichmen ools om Genoma ix sui e (www.genoma ix.de). When
P. Ma ie e al. / Da a in B ie 4 (2015) 430–436432
conside ing GO e ms associa ed o Molecula Func ion (MF) and Biological P ocess (BP), a o al o 71
GO e ms we e ound o be significan ly en iched (p- alues anges om 9.92 10
3
o 2.96 10
7
).
They we e g ouped in 18 main ca ego ies (Table 1). G oups o impo ance we e composed o eggshell
ma ix p o eins in ol ed in binding and anspo ac i i y (41 p o eins) and o 27 p o eins ela ed o
he biology o de elopmen . Addi ionally, we also epo ed p o eins in ol ed in esponse o ex e nal
s imulus (13 p o eins), in he egula ion o biological quali y (15 p o eins), exhibi ing enzyme
egula o ac i i y (13 p o eins). Also we e p esen p o eins ela ed o p o ein pos - ansla ional
olding (9 p o eins), homeos asis main enance (9 p o eins), mul i-o ganism p ocess (8 p o eins),
nu i ion and diges ion (7 p o eins), he ex acellula o ganiza ion (4 p o eins), he cell–cell adhesion
(5 p o eins), he oxido educ ase ac i i y (3 p o eins), he coagula ion p ocess (2 p o eins), he
egula ion o p o ein phospho yla ion (2 p o eins), he glyce ol e he me abolic p ocess (2 p o eins)
and he shell calcifica ion (2 p o eins).
4. Quan i a i e da ase o a ian eggshell ma ix p o eins
Two di e en me hods we e applied o disce n he ela i e abundance o he p o eins a he
di e en s ages o eggshell o ma ion, fi s ly he emPAI was calcula ed o all p o eins a each
indi idual s age and secondly, GeLC–MS/MS analyses combined wi h label ee spec al coun ing
me hod we e ca ied ou o de e mine quan i a i e alues a he ou s ages o mine aliza ion o he
216 unique p o eins.
Table 1
GO e ms significan ly en iched in he 216 iden ified eggshell ma ix p o eins.
Desc ip ions Numbe o
p o eins
Symbols
Response o ex e nal
s imulus
13 AVBD11, HSPA5, RBP4, FN1, LCN8, ALB, OC-17, CTSD, HIST1H2B7, LYZ, APOA1,
TTR, LY86
Ex acellula o ganiza ion 4 ANXA2, LOXL2, VMO1, COL2A1
Nu i ion and diges ion 7 HSPA5, ALB, CTSD, RBP4, APOA1, VTG1, VTG2
Oocy e o ma ion 3 RBP4, VMO1, TTR
P o ein pos ansla ional
olding
9 PDIA3, PPIB, QSOX1, P4HB, HSPA5, DNAJC3, HSP90B1, HSP90AA1, DNAJB6
Homeos asis
main enance
9 PDIA3, RBP4, CALB1, QSOX1, LCN8, COL2A1, P4HB, APOA1, CALM
De elopmen 27 HSPA5, SPP1, RBP4, SEMA3C, DKK3, ACTA1, ANXA2, LOXL2, CALB1, YWHAE, FN1,
ATP5B, LCN8, CDH2, TSKU, OC-17, GSN, DNAJB6, COCH, COL2A1, FST, SDF4, OC-
116, APOA1, CNTN1, CA2, TTR
Enzyme egula o ac i i y 13 HSP90AA1, ANXA2, OVM, OVST, OVAY, SERPINI1, DNAJB6, APOA1, OIH, OVAX,
TIMP2, CST3, CALM
Binding and anspo
ac i i y
41 DNAJC3, DNAJB6, CALB1, APOA1, GPC1, CNTN1, ALB, LCN8, HSPA5, SPP1, RBP4,
HSP90B1, PTN, HSP90AA1, ANXA2, LOXL2, OVM, YWHAE, RDX, ANXA5, CDH2,
GSN, COL2A1, FST, YWHAZ, HIST1H2B7, OIH, LYZ, HBG2, CDH1, TTR, CALM,
FBLN1, SDF4, SPARC, OC-17, PLOD1, HBAA, HBAD, VTG1, VTG2
Oxido educ ase ac i i y 3 PDIA3, QSOX1, P4HB
Coagula ion p ocess 2 ANXA2, ANXA5
Cell–cell adhesion 5 CDH2, DNAJB6, COL2A1, APOA1, CDH1
Ca ilage me abolism 2 LOXL2, LCN8, CDH2, COL2A1
Glyce ol e he me abolic
p ocess
2 PDIA3, P4HB
Regula ion o biological
quali y
15 PDIA3, RBP4, ANXA2, ALB, CALB1, FN1, QSOX1, RDX, LCN8, GSN, COCH, COL2A1,
P4HB, APOA1, CALM
Mul i-o ganism p ocess 8 RBP4, ALB, AVBD11, LCN8, OC-17, HIST1H2B7, LYZ, TTR
Shell calcifica ion 2 OC-17, OC-116
Regula ion o p o ein
Dephospho yla ion
2 YWHAE, CALM
P. Ma ie e al. / Da a in B ie 4 (2015) 430–436 433
In he fi s app oach, emPAI om he p o eins was calcula ed amongs he p o eins e ealed a a
pa icula s age o shell o ma ion a 5, 6, 7 and 16 h p.o. The aim was o classi y he ela i e
abundance o he p o eins wi hin one indi idual s age and o de e mine hose which we e he mos
abundan ones. EmPAI alues we e epo ed o he 216 eggshell ma ix p o eins a each s age, a
nume ical alue di e en om ze o being in oduced when he p o ein was p esen (Supplemen a y
Table 1). EmPAI alues we e also used o de e mine he numbe o p o eins p esen a a pa icula
s age (Supplemen a y Table 2). The numbe s o p o eins showing an emPAI di e en om ze o we e
91, 132, 178 and 184 a 5, 6, 7 and 16 h p.o., espec i ely.
In he second app oach, GeLC–MS/MS combined wi h label ee quan i a i e analyses based on a
spec al coun ing me hod we e used o compa e he abundances o he di e en p o eins be ween he
ou s ages o eggshell o ma ion (Supplemen a y Tables 1 and 2). One way ANOVA was pe o med on
quan i a i e alues o p o ein abundance in o de o e eal p o eins which we e a a di e en
concen a ion be ween a leas one s age ela i e o ano he one. Di e ences we e conside ed o be
s a is ically significan o p- alueo0.01. Amongs he 216 p o eins, 175 showed di e en ial
abundance acco ding o he ou s ages o shell calcifica ion. The mean abundance pe s age o shell
calcifica ion was s anda dized o each p o ein on mean alues pe s age and was calcula ed o
pe o m hie a chical clus e ing analysis. A o al o en clus e s we e highligh ed and de ailed in [1].
They we e g ouped in fi e main p o ein p ofiles associa ed wi h he di e en e en s which occu
du ing eggshell calcifica ion [1,3]: o e abundance du ing he p ima y e en s o widesp ead
deposi ion o amo phous calcium ca bona e, o aling 27 p o eins (5–6 h p.o.), o e abundance du ing
ans o ma ion o ACC in o c ys alline calci e agg ega es, o aling 13 p o eins (6 h p.o.), o e -
abundance du ing he o ma ion o la ge calci e c ys al uni s, o aling 47 p o eins (6–7 h p.o.),
o e abundance du ing apid g ow h o calci e and de elopmen o a columna s uc u e wi h
p e e en ial o ien a ion o calci e, o aling 70 p o eins (16 h p.o.). The las g oup co esponded o
o e abundance h oughou he h ee la e s ages o calcifica ion (6–7 o 16 h p.o.), o alising 18
p o eins.
5. Supplemen a y unc ional anno a ions o a ian eggshell ma ix p o eins
Po en ial unc ions o p o eins we e de e mined acco ding o he li e a u e, da a anno a ions and
unc ional domain da abase and highligh ed 77 p o eins wi h po en ial unc ions ela ed o a
mine aliza ion p ocess [1].
Beside hese p o eins, we iden ified 35 p o eins wi h po en ial an imic obial unc ions p esen a
a ious s ages o mine aliza ion (Supplemen a y Table 3). These an imic obial p o eins we e p esen a
qui e equally numbe h oughou he di e en s ages o shell o ma ion.
The mos abundan p o ein in he shell ma ix is lysozyme, which is known o hyd olyze 1–4 be a
linkages be ween N-ace yl mu amic acid and N-ace yl-
D
-glucosamine esidues in pep idoglycans o
G am-posi i e bac e ia [16]. Ano he no able p o eins is OVAX, which exhibi s an imic obial p ope ies
agains Lis e ia monocy ogenes and Salmonella en e i idis [17].A ian βde ensin 11, belonging o a ian β
de ensin amily, is a ca ionic pep ides wi h h ee s anded βshee s uc u e connec ed wi h βhai pin
loop ha p o ec agains G am-posi i e and G am-nega i e bac e ia [18].O ocledin-17, a C- ype lec in
p o ein which exhibi s an imic obial p ope ies agains Bacillus sub ilis,S aphylococcus au eus and
Pseudomonas ae uginosa is also a majo eggshell p o ein [19].
Ano he g oup is cons i u ed o an imic obial p o eins by dep i ing bac e ia o essen ial nu ien s.
O o ans e in, i ellogenin-1 and 2 and MFI2 a e an imic obial by hei capaci y o chela e i on ions,
essen ial o bac e ial g ow h [20].GC, a g oup-specific componen and ibofla in p o ein bind
i amins which a e dep i ing bac e ia om i amin D.
We epo o ocalyxin-36,BPIFCB and o oglobulinG2 ype AA, which belong o he LBP/BPI/PLUNC
amily known o bind o he lipid A po ion o lipopolysaccha ide cell wall in G am-nega i e bac e ia
leading o he dea h o bac e ia [21].
The s udy also e ealed p o ease inhibi o s which can be po en ially an imic obial by hei abili y
o inhibi p o eases sec e ed by some bac e ia. O oinhibi o ,o omucoid,SPARC, ollis a in, ollis a in-like
1and IGFBP7 exhibi Kazal like p o ease inhibi o domains. Mo eo e , o oinhibi o was showed o
P. Ma ie e al. / Da a in B ie 4 (2015) 430–436434

inhibi Bacillus hu ingiensis g ow h [22].O ocalyxin-32 p esen s homology wi h la exin, a
ca boxypep idase inhibi o , and has been shown o inhibi B. sub ilis g ow h [23].SERPIND1,SERPINF2,
SERPINE2,SERPING1 and SERPINI1 belong o SERPIN amily, a known amily o se ine p o ease
inhibi o s [16].O ocalyxin-25 con ains wo inhibi o p o ease domains, a WAP and a Kuni z-like
domain [24].Cys a in C is a cys eine p o ease inhibi o , and o os a in is known o inhibi all ou
classes o p o eases [16].
Finally ou s udy e ealed se e al agmen o immunoglobulin (Ig-ligh -2,Ig-gamma-hea y,Ig-
lambda-1,Ig-lambda-2,Ig-ligh -1,Ig-mu-hea y and Ig-alpha-hea y).
Finally, ou o he 175 eggshell ma ix p o eins showing significan a ia ion o abundance
acco ding o he ou s ages o shell calcifica ion, 81 could no be asc ibed o any o hese unc ional
g oups and we e classified as “o he o unknown ole”(Supplemen a y Table 4).
Conflic o in e es
Au ho s claim no conflic o in e es .
Acknowledgmen s
This esea ch was unded by he F ench Na ional Resea ch Agency ANR (ANR-13-BSV6-0007-01,
ANR-13-BSV6-0007-02 and ANR-13-BSV6-0007-05). The high esolu ion mass spec ome e was
financed (SMHART p ojec , 35069) by he Eu opean Regional De elopmen Fund (ERDF), he Conseil
Régional du Cen e, he F ench Na ional Ins i u e o Ag icul u al Resea ch (INRA) and he F ench
Na ional Ins i u e o Heal h and Medical Resea ch (Inse m). ARN acknowledges unding h ough
G an s CGL2011-25906 (Minis e io de Economia, Spain).
The au ho s a e g a e ul o he expe imen al uni s (UE-PEAT) o ca e o bi ds, and o he geno oul
bioin o ma ics pla o m Toulouse Midi-Py enees o p o iding help and/o compu ing and/o s o age
esou ces.
Appendix A. Suppo ing in o ma ion
Supplemen a y da a associa ed wi h his a icle can be ound in he online e sion a h p://dx.doi.
o g/10.1016/j.dib.2015.06.019.
Re e ences
[1] P. Ma ie, V. Labas, A. B ionne, G. Ha ichaux, C. Henneque -An ie , A.B. Rod iguez-Na a o, e al., Quan i a i e p o eomics
p o ides new insigh s in o chicken eggshell ma ix p o ein unc ions du ing he p ima y e en s o mine aliza ion and he
ac i e calcifica ion phase, J. P o eomics 126 (2015) 140–154.
[2] J.A. Vizcaino, E.W. Deu sch, R. Wang, A. Cso das, F. Reisinge , D. Rios, e al., P o eomeXchange p o ides globally coo dina ed
p o eomics da a submission and dissemina ion, Na . Bio echnol. 32 (2014) 223–226.
[3] A.B. Rod iguez-Na a o, P. Ma ie, Y. Nys, M.T. Hincke, J. Gau on, Amo phous calcium ca bona e con ols a ian eggshell
mine aliza ion: a new pa adigm o unde s anding apid eggshell calcifica ion, J. S uc . Biol. 190 (2015) 291–303.
[4] V. Labas, I. G asseau, K. Cahie , A. Ga ga os, G. Ha ichaux, A.P. Teixei a-Gomes, e al., Quali a i e and quan i a i e
pep idomic and p o eomic app oaches o pheno yping chicken semen, J. P o eomics 112 (2015) 313–335.
[5] A. Kelle , A.I. Nes izhskii, E. Kolke , R. Aebe sold, Empi ical s a is ical model o es ima e he accu acy o pep ide
iden ifica ions made by MS/MS and da abase sea ch, Anal. Chem. 74 (2002) 5383–5392.
[6] A.I. Nes izhskii, A. Kelle , E. Kolke , R. Aebe sold, A s a is ical model o iden i ying p o eins by andem mass spec ome y,
Anal. Chem. 75 (2003) 4646–4658.
[7] F. Sie e s, A. Wilm, D. Dineen, T.J. Gibson, K. Ka plus, W.Z. Li, e al., Fas , scalable gene a ion o high-quali y p o ein mul iple
sequence alignmen s using Clus al Omega, Mol. Sys . Biol. (2011) 7.
[8] C. Camacho, G. Coulou is, V. A agyan, N. Ma, J. Papadopoulos, K. Beale , e al., BLAST plus: a chi ec u e and applica ions,
BMC Bioin o m. (2009) 10.
[9] K. Mann, B. Macek, J.V. Olsen, P o eomic analysis o he acid-soluble o ganic ma ix o he chicken calcified eggshell laye ,
P o eomics 6 (2006) 3801–3810.
P. Ma ie e al. / Da a in B ie 4 (2015) 430–436 435
[10] K. Mann, J.V. Olsen, B. Macek, F. Gnad, M. Mann, Phosphop o eins o he chicken eggshell calcified laye , P o eomics 7
(2007) 106–115.
[11] I. Miksik, J. Cha a o a, A. Eckha d , Z. Deyl, Insoluble eggshell ma ix p o eins – hei pep ide mapping and pa ial
cha ac e iza ion by capilla y elec opho esis and high-pe o mance liquid ch oma og aphy, Elec opho esis 24 (2003)
843–852.
[12] I. Miksik, A. Eckha d , P. Sedlako a, K. Mikuliko a, P o eins o insoluble ma ix o A ian (Gallus Gallus) eggshell, Connec .
Tissue Res. 48 (2007) 1–8.
[13] I. Miksik, P. Sedlako a, K. Lacino a, S. Pa a idis, A. Eckha d , De e mina ion o insoluble a ian eggshell ma ix p o eins,
Anal. Bioanal. Chem. 397 (2010) 205–214.
[14] M. Rose-Ma el, J.W. Du, M.T. Hincke, P o eomic analysis p o ides new insigh in o he chicken eggshell cu icle,
J. P o eomics 75 (2012) 2697–2706.
[15] C.J. Sun, G.Y. Xu, N. Yang, Di e en ial label- ee quan i a i e p o eomic analysis o a ian eggshell ma ix and u e ine fluid
p o eins associa ed wi h eggshell mechanical p ope y, P o eomics 13 (2013) 3523–3536.
[16] S. Réhaul -Godbe , V. He é, J. Gau on, C. Cabau, Y. Nys, Molecules in ol ed in chemical de ence o he chicken egg, in:
Y. Nys, M. Bain, F. Van Imme seel (Eds.), Imp o ing he Sa e y and Quali y o Eggs and egg P oduc s, Woodhead Publishing
limi ed, Camb idge 2011, pp. 183–208.
[17] S. Rehaul -Godbe , V. Labas, E. Helloin, V. He e-G epine , C. Slugocki, M. Be ges, e al., O albumin- ela ed p o ein X is a
hepa in-binding o -se pin exhibi ing an imic obial ac i i ies, J. Biol. Chem. 288 (2013) 17285–17295.
[18] V. He e-G epine , S. Rehaul -Godbe , V. Labas, T. Magallon, C. De ache, M. La e gne, e al., Pu ifica ion and
cha ac e iza ion o a ian be a-de ensin 11, an an imic obial pep ide o he hen egg, An imic ob. Agen s Chemo he . 54
(2010) 4401–4408.
[19] O. Wellman-Labadie, J. Picman, M.T. Hincke, An imic obial ac i i y o cu icle and ou e eggshell p o ein ex ac s om h ee
species o domes ic bi ds, B . Poul . Sci. 49 (2008) 133–143.
[20] J.A. Ga ibaldi, Role o mic obial i on anspo compounds in bac e ial spoilage o eggs, Appl. Mic obiol. 20 (1970) 558–560.
[21] J. Gau on, S. Rehaul -Godbe , G. Pascal, Y. Nys, M.T. Hincke, O ocalyxin-36 and o he LBP/BPI/PLUNC-like p o eins as
molecula ac o s o he mechanisms o he a ian egg na u al de ences, Biochem. Soc. T ans. 39 (2011) 971–976.
[22] M. Bou in, J. Gau on, M. Be ges, S. A ucci, G. Le Blay, V. Labas, e al., An imic obial po en ial o egg yolk o oinhibi o , a
mul idomain kazal-like inhibi o o chicken egg, J. Ag ic. Food Chem. 59 (2011) 12368–12374.
[23] J. Xing, O. Wellman-Labadie, J. Gau on, M.T. Hincke, Recombinan eggshell o ocalyxin-32: exp ession, pu ifica ion and
biological ac i i y o he glu a hione S- ans e ase usion p o ein, Comp. Biochem. Pysiol. B Biochem. Mol. Biol. 147 (2007)
172–177.
[24] J. Gau on, Y. Nys, Func ion o eggshell ma ix p o eins, in: R. Huopalah i, R. Lopez-Fandino, M. An on, R. Schade (Eds.),
Bioac i e Egg Compounds, Sp inge -Ve lagGe many 2007, pp. 109–115.
P. Ma ie e al. / Da a in B ie 4 (2015) 430–436436