Exploring the interaction sites in glucose and galactose using phenol as a probe

This jou nal is © he Owne Socie ies 2023 Phys. Chem. Chem. Phys., 2023, 25, 7205–7212 | 7205
Ci e his: Phys. Chem. Chem. Phys.,
2023, 25, 7205
Explo ing he in e ac ion si es in glucose and
galac ose using phenol as a p obe†
Pau
´l Pinillos, ‡Ande Cami uaga, ‡Fe nando To es-He na
´ndez,
F ancisco J. Bas e echea, Imanol Usabiaga * and Jose
´A. Fe na
´ndez *
Suga s, oge he wi h amino acids and nucleobases, a e he undamen al building blocks o a cell. They
a e in ol ed in many undamen al p ocesses and hey especially play ele an oles as pa o he
immune sys em. The la e is connec ed o hei abili y o es ablish a collec ion o in e molecula
in e ac ions, depending on he posi ion o hei hyd oxyl g oups. He e we explo e how he posi ion o
he OH in C4, he anome ic con o ma ion and he na u e subs i uen affec he in e ac ion wi h phenol,
which se es as a p obe o he p e e ed si e o he in e ac ion. Using mass- esol ed exci a ion
spec oscopy and densi y unc ional calcula ions, we un a el he s uc u e o he dime s and compa e
hei con o ma ion wi h hose ound o simila sys ems. The main conclusion is ha he hyd oxyme hyl
g oup has a e y s ong in luence, guiding he whole agg ega ion p ocess and ha he posi ion o he
subs i uen in C4 has a s onge in luence on he inal s uc u e o he dime han he anome ic
con o ma ion.
In oduc ion
Suga s, oge he wi h nucleobases, amino acids and lipids, a e
he building blocks o a cell. They play undamen al oles om
ene gy s o age o cons i uen molecules o some biopolyme s,
such as s a ch and cellulose.
1,2
One o he mos ascina ing
asks hey a e in ol ed in is he immune unc ion. Combining
se e al suga uni s, cells c ea e polysaccha ides ha p esen
unique con o ma ional and in e ac ional landscapes, de ined
by he na u e o hei monosaccha ide componen s.
3
The
co esponding ecep o s in he immune cells a e able o sense,
p obe and ecognize such con igu a ions and de e mine i he
cell is iend o oe. I is su p ising how he immune sys em is
able o dis inguish e en he smalles modi ica ion: he axial/
equa o ial posi ion o a hyd oxyl g oup in one o he mono-
saccha ide cons i uen s.
4
In pa , his abili y elies on a kind o
ampli ica ion effec p oduced by he hyd ogen bond ne wo ks
ha ex end along he whole glycan. Modi ica ion o he posi-
ion o a gi en hyd oxyl g oup om axial o equa o ial o ice
e sa al e s such a ne wo k. This acili a ed he co esponding
ecep o o de ec he s uc u al diffe ences.
5,6
To unde s and he whole ecogni ion p ocess, in o ma ion
ega ding wo aspec s o he sys em is essen ial: a deep
knowledge o he s uc u e o mono- and poly-saccha ides
and o he in e ac ions ha hey can es ablish wi h o he
molecules, such as o example, he la e al chains o amino
acids. The e a e many esea ch g oups con ibu ing o inc easing
he knowledge on he s uc u e o saccha ides. Since he pionee -
ing wo ks o Simons’ g oup using lase spec oscopy,
7
he main
ad ances ha e come om he high esol ing powe o mic owa e
spec oscopy.
8–15
On he o he hand, he la ge size o he agg ega es be ween
monosaccha ides and o he molecules usually makes he use o
o he echniques o ex ac s uc u al in o ma ion necessa y.
Two echniques a e commonly used: NMR and mass- esol ed
spec oscopy in je s. While he o me gi es impo an in o ma-
ion ega ding he beha iou o he saccha ides in solu ion,
16
he
la e enables he cha ac e iza ion o he mos s able s uc u es
in he absence o ex e nal pe u ba ions.
17
Ye , such s udies a e
no easy: i s , he saccha ides usually do no con ain a ch omo-
pho e and he e o e, i is necessa y o modi y he molecule o
include an a oma ic subs i uen . This is usually done in he
anome ic ca bon, o a oid spon aneous a/bisome iza ion. Ce -
ainly, monosaccha ides p esen a linea o m, especially in
solu ion, which allow hem o in e con e be ween anome s.
Second, suga s canno be ans e ed o he gas phase by simple
wa ming. They usually equi e a deso p ion sys em, which
in oduces addi ional noise and limi s he maximum signal
in ensi y achie able. Signals become less s able and he spec a
noisie , complica ing hei acquisi ion and in e p e a ion. This
is p obably one o he easons why no many g oups ha e
engaged in he s udy o he s uc u e o suga agg ega es.
Depa men o Physical Chemis y, Fac. o Science and Technology, Uni e si y o he
Basque Coun y (UPV/EHU), Ba io Sa iena s/n, 48940, Spain.
E-mail: josea. e nan[email p o ec ed], i.usabiag[email p o ec ed]
†Elec onic supplemen a y in o ma ion (ESI) a ailable: Addi ional compu ed
s uc u es and p edic ed IR spec a. See DOI: h ps://doi.o g/10.1039/d2cp06036a
‡These wo au ho s con ibu ed equally o his wo k.
Recei ed 27 h Decembe 2022,
Accep ed 6 h Feb ua y 2023
DOI: 10.1039/d2cp06036a
sc.li/pccp
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Ou g oup has con ibu ed o his ield wi h he s udy o he
agg ega ion p e e ences o glucose (Glc), galac ose (Gal) and
se e al de i a i es wi h a collec ion o molecules: om suga
de i a i es o d ugs, using se e al mass- esol ed lase spec o-
scopic echniques.
5,6,18–20
He e, we ex end hose p e ious s udies o he agg ega ion
p e e ences o Gal de i a i es wi h phenol (PhOH, Scheme 1)
and compa e he esul s ob ained wi h hose ound o he
agg ega ion o Glc de i a i es wi h PhOH. As can be seen, Glc
and Gal only diffe in he posi ion o he hyd oxyl g oup
a ached o C4. Such a small s uc u al diffe ence p oduces a
e-accommoda ion o he es o OH g oups in he molecule
and a change in he o ien a ion o he in amolecula hyd ogen
bond ne wo k, also modi ying he way in which he suga uni
in e ac s wi h o he molecules.
Two de i a i es o each monosaccha ide we e included in
his s udy: wi h an –OMe and wi h an –OPhe g oup in he
anome ic ca bon, ei he in he ao in he bcon igu a ion. The
compa ison be ween he s uc u es o he agg ega es wi h each
o he wo subs i uen s allowed us o e alua e he endency o
he suga owa ds s acking, a ype o in e ac ion necessa y o
unde s and he agg ega ion o nucleobases. On he o he hand,
pas s udies demons a ed ha he sub le s uc u al a ia ion
in oduced by he a/bcon o ma ion o he anome ic subs i uen
esul ed in e y di e en agg ega ion s uc u es and in e ac ion
ene gy alues.
6
The e o e, we also analyse he e he anome ic
e ec on he s uc u e o he agg ega es.
Me hods
Expe imen al
The expe imen al se up has been ex ensi ely desc ibed in
p e ious publica ions and he e o e only he mos ele an
aspec s will be offe ed he e. The sys em is buil a ound a linea
ime o ligh mass spec ome e ha ac s as a mass-disc iminan
de ec o . The sample, consis ing o a mix u e o suga de i a-
i es, phenol and ca bon nano ubes (MWCNT, Cheap ubes
Inc.), was a ached o a cylind ical sample holde as desc ibed
in Usabiaga e al.,
19
and placed on one side a he exi o a
pulsed al e (Gene al Val e Se ies 9). In his way, he sample
deso bed by he IR pho ons om he abla ion lase (Nd/YAG
Quan el B illian B, 1064 nm 0.5–1 mJ pe pulse ocused on he
sample) was picked by he expanding gas (A 12 ba , 99.999%
pu i y, P axai ) a each al e opening, and cooled by he sub-
sequen expansion. Unde such condi ions, agg ega es o med
and a elled as a dense molecula beam ha was in e oga ed
using a combina ion o pulsed ns lase s (UV: Quan el Q-Scan,
Couma in 540 A, 500 mJ pe pulse and IR: Lase Vision OPO/A,
B7 mJ pe pulse). One-colo REMPI spec oscopy was used o
ob ain signal om each molecula agg ega e, while IR/UV
double esonance allowed us o ex ac con o me -selec i e s uc-
u al in o ma ion o compa e wi h he compu a ional p edic ions
and o e a sound assignmen o each species de ec ed. Pu i y o
he compounds s udied: phenyl-b,D-glucopy anoside (b-PhGlc),
97%; phenyl-b,D-galac opy anoside (b-PhGal): 98%; me hyl-b,D-
glucopy anoside (b-MeGlc): Z99%; me hyl-b,D-galac opy-
anoside (b-MeGal): Z98%; me hyl-a,D-glucopy anoside (a-
MeGlc): Z99%; and me hyl-a,D-galac opy anoside (a-MeGal): Z
99%. All o hem we e pu chased om Sigma-Ald ich (Mad id,
Spain).
Compu a ions
The compu a ional p ocedu e has been de ailed in p e ious
publica ions.
21
B ie ly, i was di ided in o h ee s ages. Fi s ,
se e al o ce ields (MMFFs,
22
AMBER
23
and OPLS3e
24
) we e
used o explo e he con o ma ional landscape o each dime .
The (usually) housands o s uc u es gene a ed his way we e
g ouped in o amilies wi h a simila balance o in e ac ions and
which e y likely p esen shallow ba ie s o isome iza ion.
The lowes ene gy membe o each amily, oge he wi h some
o he selec ed s uc u es we e subjec ed o ull op imiza ion
using Densi y Func ional Theo y (DFT) wi h M06-2X and
B3LYP-D3 unc ionals and 6-311++G(d,p) basis unc ions:
M06-2X/6-311++G(d,p) and B3LYP-D3/6-311++G(d,p).
Once he inal se o s uc u es was ob ained, hei IR
spec a we e simula ed using he no mal modes gene a ed in
he co esponding calcula ion and an algo i hm ha akes in o
accoun he anha monici y by in oducing a pa ame e (0.953
o CHs and 0.9385 o OHs o he calcula ions a he M06-2X/
6-311++G(d,p) le el and 0.968 o CHs and 0.9535 o OHs o he
calcula ions a he B3LYP-D3/6-311++G(d,p) le el espec i ely).
The b oadening in he OH s e ching ansi ions in oduced by
he o ma ion o he hyd ogen bonds was also included using a
polynomial exp ession:
Dn= 1 + (80 (3600 X
F eq
)/(3600–3200))
whe e Dnis he b oadening o a gi en ansi ion, X
F eq
is he
posi ion o he ansi ion in cm
1
and he es a e empi ical
pa ame e s ob ained om i ing o he wid h o he lines in he
spec a o a collec ion o sys ems.
Scheme 1 S uc u es o phenol, phenyl-b,D-glucopy anose, me hyl-b,D-
glucopy anose, me hyl-a,D-glucopy anose and hei galac ose analogues.
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Finally, he lase bandwid h was also in oduced by con ol-
ing he spec um (Lo en zian unc ion o each equency)
wi h a Gaussian unc ion o 5 cm
1
FWHM.
Resul s
Elec onic exci a ion spec oscopy o he agg ega es
Fig. 1 summa izes he 1-colo REMPI spec a o he species s udied
in his wo k. The spec a o he monome s a e well- esol ed
abso p ions ha con ain con ibu ions om se e al con o me s.
The spec um o phenol, o example, has al eady been epo ed by
se e al au ho s
25–29
and is well- esol ed, p esen ing some ib onic
ac i i y. The spec um p esen ed in Fig. 1 was eco ded using he
abla ion sys em and he e o e is no as cold as hose eco ded using
o he sou ces, bu s ill, he 0
0
0
ansi ion appea s a 36 349 cm
1
,in
good ag eemen wi h published da a.
26–32
Ve y close in ene gy bu sligh ly o highe wa enumbe s, he
spec um o PhGlc and PhGal appea . They a e e y simila ,
wi h well-de ined ansi ions ha hide he con ibu ion om
se e al con o ma ional isome s.
The con o ma ional a iabili y o hexoses a ises om he
lexibili y o he hyd oxyme hyl g oup and he o ien a ion o
he in amolecula hyd ogen bond ne wo k: ei he clockwise o
an iclockwise. P e ious s udies using mass- esol ed exci a ion
spec oscopy (MRES) al eady iden i ied h ee isome s o PhGlc,
diffe ing in he ela i e o ien a ion o he hyd oxyme hyl
g oup,
33
and ano he h ee isome s o PhGal, al hough in he
la e , one o he isome s is based in a diffe en o ien a ion o
he hyd ogen bond (hbond) ibbon.
34
In s a k con as o he spec a o he monome s, hose o
he dime s a e uns uc u ed abso p ions, wi h a e y limi ed
numbe o disc e e ea u es. The b oadening seems o be
connec ed o he na u e o he agg ega ion pa ne . P e ious
epo s on he MRES o suga uni s and on hei agg ega es
demons a ed ha e en disaccha ides agged wi h a ch omo-
pho e p esen disc e e spec a.
3
A backg ound abso p ion s a s
appea ing in he elec onic spec a o he monohyd a es
35
and
becomes mo e e iden in he dihyd a es
36,37
o become uns uc-
u ed abso p ions in he dime s wi h la ge molecules.
38
Howe e , he loss o ib onic s uc u e does no hampe eco ding
a clean and disc e e mass- esol ed IR spec um, as demons a ed
p e iously.
38
Mass- esol ed IR spec oscopy
Fig. 2 shows a compa ison be ween he mass- esol ed IR
spec a o he dime s s udied in his wo k and he p edic ions
buil using he no mal mode analysis a he M06-2x/
6-311++G(d,p) le el. The es o calcula ions may be ound in
he ESI.†Thecompa isonin he igu eshowsanexcellen
ag eemen be ween simula ions and expe imen al da a. Such an
ag eemen is magni ied by he compa ison wi h he p edic ions a
he B3LYP-D3/6-311++G(d,p) le el (ESI†). Al hough compu a ions
a he la e le el also ep oduce he gene al shape o he spec a,
he e is a la ge disc epancy in he posi ion o he bands,
especially a he high wa enumbe -end o he spec a, whe e he
ib a ions a e less anha monic and he e o e, hey a e supposed
o be be e ep oduced by he calcula ions.
Sligh ly la ge diffe ences be ween expe imen and p edic ions
we e ound o b-MeGlcPhOH, o which he calcula ions
p edic a smalle shi o he OH s e ching o he phenolic
hyd oxyl g oup. Appa en ly, he expe imen al s uc u e p esen s a
s onge hbond han p edic ed. The simula ion o he second
mos s able isome (op ion 2 in he igu e) be e ep oduces he
posi ion o he O
Ph
Hs e ch(Fig.S5,ESI†),bu a heexpenseo a
wo se ag eemen wi h he bands a ound 3600 cm
1
.Thosebands
a e due o he s e ching o he OH g oups in ol ed in mild
in e ac ions and he e o e, p esen he lowes anha monici y.
In consequence, hey should be desc ibed mo e accu a ely by
he compu a ional me hods used. In conclusion, we p e e o
main ain he assignmen o he mos s able s uc u e.
In e es ingly, a single isome was ound o each agg ega e,
excep o a-MeGlcPhOH, o which he expe imen al ace
eco ded p obing diffe en wa enumbe s poin o he exis ence
o a leas wo isome s. De e mining he exac numbe o
isome s o hese sys ems is no an easy ask, as he employmen
o double esonance echniques such as UV/UV hole bu ning is
no possible, due o he un esol ed na u e o he elec onic
spec a. Howe e , he IR spec a we e eco ded a se e al wa e-
leng hs, always ob aining he same esul s, excep o he abo e-
men ioned case o a-MeGlcPhOH.
Assigned s uc u es
A summa y o he p oposed assignmen s o he expe imen al
spec a may be ound in Fig. 3, while he comple e se o
compu ed s uc u es is collec ed in he ESI.†
Fig. 1 1-colo REMPI spec a o phenol, b-PhGlc, b-PhGal, a-MeGalPhOH,
b-MeGlcPhOH, b-MeGalPhOH, b-MeGlcPhOH, b-PhGalPhOH and
b-PhGlcPhOH. 0
0
0
ansi ions and hose wa enumbe s used o eco d he IR/
UV aces a e indica ed.
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The e is a clea p e e ence o phenol o ac as a p o on
dono o he suga uni , independen ly o he anome and/o
he anome ic subs i uen . Also in e es ing is he s ong p o-
pensi y o he in e ac ion o PhOH wi h he hyd oxyme hyl
g oup. Al hough he calcula ions demons a e ha se e al
con o me s wi h e y simila s abili y a e possible, i seems
ha he molecules always ind a low-ene gy-ba ie pa h o he
global minimum, excep o a-MeGlcPhOH.
In e es ingly, he axial/equa o ial posi ion o he hyd oxyl
moie y in C4 also modi ies he a ack angle o he phenol,
h ough he al e a ion o he posi ion o he hyd oxyme hyl
g oup. Such modi ica ion is ampli ied and ansmi ed o he
subs i uen , mainly because, in all he sys ems s udied, he e is
a clea p e e ence o maximizing he O
Ph
HO in e ac ion
while he CHpcon ac s appea as seconda y in e ac ions.
This is clea ly seen compa ing b-MeGlc and b-MeGal agg e-
ga es: in bo h cases PhOH ‘‘a acks’’ one o he lone pai s o he
Fig. 2 Mass- esol ed IR spec a o PhOH agg ega es wi h b-PhGlc,
b-PhGal, b-MeGlc, b-MeGal, a-MeGlc and a-MeGal, oge he wi h he
simula ed spec a o he isome s o which hey we e assigned (in blue),
buil using he no mal mode analysis compu ed a he M06-2x/
6-311++G(d,p) le el, and he p ocedu e desc ibed in he Me hods sec ion.
Fig. 3 Assigned s uc u es o PhOH agg ega es wi h b-PhGlc, b-PhGal,
b-MeGlc, b-MeGal, a-MeGlc and a-MeGal, compu ed a he M06-2x/
6-311++G(d,p) le el. Numbe s in b acke s a e he ela i e ene gy and he
binding ene gy in kJ mol
1
calcula ed a he speci ied empe a u es. The
es o he compu ed s uc u es may be ound in he ESI.†
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O6H. Howe e , he la e is below he plane o he suga ing in
b-MeGlc o in e ac wi h O4H and abo e he plane in b-MeGal.
This diffe ence is mainly a o a ion o he C4C5C6O6 dihed al
angle, om 71 o 71 deg ees, and esul s in a comple ely
diffe en o ien a ion o he PhOH, which in he o me in e ac s
wi h he C
ing
H a oms and wi h he C
b
H
3
in he la e . A simila
shi in he posi ion o he PhOH was obse ed o a-MeGlc/Gal.
Discussion
In gene al, he assignmen in all dime s s udied he e is e y
simila and co esponds o he same molecula amily, in
which phenol ac s as a p o on-dono o he O6 o suga ( he
amily in blue). A second amily o s uc u es, ma ked in g een
in Fig. 2 and 3, in which O4H om he suga o ms a seconda y
hyd ogen bond wi h he phenolic oxygen, seems o be p esen
a leas o one o he dime s and i is in gene al e y close in
s abili y o he global minimum. This seconda y amily has he
peculia i y ha he binding ene gy is maximized, a he expense
o a educ ion on he s abili y o he suga uni .
In luence o he anome ic con o ma ion and subs i uen on he
agg ega ion p ocess
Apa om he diffe ence in he posi ion o he subs i uen in C4,
he monome s s udied p esen wo a ia ions: he a/banome ic
con o ma ion and he subs i uen a ha posi ion: ei he a MeO–
o a PhO– g oup. In p inciple, one would expec he phenyl-
subs i u ed monosaccha ides o gi e ise o s onge binding
ene gies, as i gi es he molecules he chance o es ablish a
collec ion o addi ional in e ac ions (pp s acking o CH–p). One
would expec hose sys ems wi h he PhO– subs i uen o p esen
a s onge binding ene gy, simply because he PhO– g oup is
la ge and he e o e, mo e pola izable. Ce ainly, he e is a
subs an ial di e ence in he posi ion o PhOH espec o he
suga uni , depending on he anome ic subs i uen . Howe e , he
calcula ions p edic simila binding ene gy alues o bo h
se s o species, o e en lowe o he phenolic-subs i u ed ones
(Table 1).
The anome ic con o ma ion does no seem o ha e a s ong
in luence on he binding ene gy. The la ges changes in hese
alues seem o be ela ed o he con o ma ion o he subs i uen
in C4, a leas o he me hyl-subs i u ed species. I is clea om
he able ha he a-/b-MeGal species p esen B10% highe
binding ene gy alues han hei homologous Glc species.
The shi in he posi ion o he OH s e ching band is usually
a good indica o o he s eng h o a hyd ogen bond. As can be
seen in Fig. 4, wo diffe en co ela ions may be es ablished
be ween hese wo pa ame e s: one o a-/b-MeGal plus b-MeGlc
and a second linea co ela ion wi h a smalle slope o he es
o he species plus b-MeGlc. The second isome assigned o
a-MeGlc does no i well in his gene al pic u e, al hough i is
close o he alues o a-/b-MeGal. The s eepe slope o he ed
i , means ha he inc ease in binding ene gy p oduced a
smalle shi in he posi ion o he s e ching o he phenolic
OH, indica ing ha he ex a ene gy may come om o he
in e ac ions ins ead o he ein o cemen o he in e molecula
hyd ogen bond. Ce ainly, when he s eng h o he in e ac ions
is compu ed using he bond c i ical poin s, all he alues i o a
single s aigh line (Fig. 5).
Compa ison wi h simila sys ems: he in luence o he s acking
in e ac ion
Mos s udies ha e cen ed on ackling he s uc u e o suga –
wa e o suga –a oma ic agg ega es. The eason behind he
o me s udies is clea : wa e is he main cons i uen o he
biological en i onmen . On he o he hand, suga –a oma ic
in e ac ions a e o high ele ance in biological en i onmen s:
mos o he suga –p o ein in e ac ions in ol e con ac s wi h
a oma ic amino acids.
39,40
Compa ison o he esul s ob ained in he p esen wo k wi h
hose om he monohyd a es may help us unde s and he
in luence o he in e ac ions due o dispe si e o ces in he
inal s uc u e o he agg ega es. Howe e , one o he p oblems
encoun e ed o such compa a i e s udy is ha some o he
published wo ks a e al eady close o 20 yea s old and ask o a
e-in es iga ion, especially due o he compu a ional le els used
o in e p e he expe imen al esul s, which we e no as accu a e
as he s a e-o - he-a unc ional. Ce ainly, he ca bohyd a e–
a oma ic in e ac ion is conside ed a es o DFT calcula ions,
because o he delica e balance be ween he in e ac ions in ol-
ing OH and CH g oups.
41
Ne e heless, he e is a gene al ag eemen in ha wa e
inse s in he weakes in amolecula hbond o he suga .
42
In
Glc and mannose, wa e ends o in e ac wi h O4H as a p o on-
Table 1 Dissocia ion ene gy alues a 0 K and posi ion o he s e ching
ib a ions o he dime s s udied in his wo k
Dime
Binding ene gy
0 K (kJ mol
1
)
s(PhOH)/
cm
1
s(OH)
suga
/cm
1
b-PhGlcPhOH 44.3 3390 3595, 3645
b-PhGalPhOH 45.6 3360 3600, 3606, 3632, 3645
b-MeGlcPhOH 43.0 3420 3529, 3642
b-MeGalPhOH 48.4 3405 3548, 3596, 3636
a-MeGlcPhOH I 44.2 3380 3562, 3594, 3634
a-MeGlcPhOH II 50.3 3415 3521, 3618, 3643
a-MeGalPhOH 50.9 3390 3541, 3594, 3616
Fig. 4 Binding ene gy o he dime s s udied in his wo k s. posi ion o he
s e ching o he phenolic OH.
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accep o and, a he same ime, o dona e o O6, o ming an
eigh -membe ed ing.
33,35,43,44
Addi ional s udies on Glc-wa e
ound a seconda y isome wi h he wa e molecule apped
be ween he O6H and he ing’s oxygen a om,
34
which is
p ecisely he p e e ed con o ma ion o Gal monohyd a es.
36
Thus, in he case o suga –wa e in e ac ion, he p e e ed
sol a ion si e changes wi h he monosaccha ide, while in he
case o he dime s wi h phenol, he global minimum always has
he phenol molecule ac ing as p o on-dono o he sugga ’s
O6H. This is ue a leas o he wo monosaccha ides s udied
in his wo k. This may be he esul o se e al ac o s: phenol is
a be e p o on-dono han wa e , and he e o e, i may esul
mo e e icien o he sys em o place he phenol molecule a
he ail o he hyd ogen-bond ne wo k. I is also a poo e
hyd ogen bond accep o , and he e o e, i may no be able o
inse be ween he O4H and O6, as wa e does in i s in e ac ion
wi h Glc. Fu he mo e, adop ing he posi ion shown in Fig. 3, i
may be able o maximize o he seconda y in e ac ions.
To e alua e he ela i e weigh o each o hese ac o s i
would be in e es ing o ex end he s udies on phenol–suga
dime s o o he monosaccha ides, in which wa e o ms wha
has been e med as ‘‘inse ion s uc u es’’: isome s in which
wa e inse s be ween O3H and O4H o be ween O2H and O3H,
b eaking an in amolecula hyd ogen bond.
43,45–48
Rega ding he suga –a oma ic in e ac ion, se e al s udies
demons a ed ha he op imal in e ac ion akes place wi h he
hyd ogen a oms o C3, 4 and 5.
41,49–54
These in e ac ions a e
maximized in hose monosaccha ides in which all he OHs a e on
onesideo heplaneo he ing,suchasin ucose.
53
In e es ingly,
a compu a ional s udy on ucose–phenol dime using CCSD(T)
and ex apola ing o he basis se limi , p edic ed a s uc u e o
he global minimum subs an ially di e en om hose in Fig. 3,
wi h he phenol o ming pa o an O5HO
PhOH
HO
ing
hyd o-
gen bond ne wo k and he a oma ic ing poin ing away om he
suga . Con e sely, in all he sys ems s udied he e, phenol ac s as a
p o on-dono o he hyd oxyme hyl g oup. Assuming ha he
au ho s did no o e look any isome , an always p esen isk in
his kind o complex sys em wi h a mul i ude o local minima, he
eason o his di e ence wi h he dime s s udied in his wo k
may be he absence o a hyd oxyme hyl g oup in ucose. This
unc ional g oup may comple ely modi y he con o ma ional
landscape. Ce ainly, he global minimum o ucose–wa e p e-
sen s an inse ion s uc u e, as men ioned abo e,
46
di e ing om
he s uc u es ound o Glc/Gal-wa e .
33,37
Compa ison be ween he s uc u es in Fig. 3 and hose ound
o monosaccha ide–a oma ic (a oma ic = benzene, oluene o
indole) agg ega es
40,41,49–54
shows ha he hyd oxyl g oup is a
game change . While in he o me sys ems he a oma ic always
lies pa allel o he saccha ide ing, phenol–suga in e ac ion is
guided by he op imiza ion o he hyd ogen bond. Only i he
o ien a ion o he O6H allows i , phenol can adjus he posi ion
o i s a oma ic ing o also maximize he in e ac ion wi h he
alipha ic hyd ogens o he monosaccha ide, such as in a-/b-
GlcPhOH. I is also wo h men ioning ha he in e ac ion
wi h phenol ixes he o ien a ion o he hyd oxyme hyl g oup in a
single posi ion, collapsing he collec ion o o ame s obse ed
o he monosaccha ides in o a single species. This is also in line
wi h he obse a ion o se e al au ho s, which connec in e -
ac ion wi h he sol en wi h a simpli ica ion o he con o ma-
ional landscape o he suga uni s.
36
Conclusions
The dime s o a-/b-MeGlc, a-/b-PhGlc, a-/b-MeGal and a-/b-
PhGal wi h phenol ha e been cha ac e ized by a combina ion
o lase spec oscopy in je s and DFT calcula ions. In all cases,
he p e e ed in e ac ion si e is he hyd oxyme hyl g oup, wi h
phenol ac ing as he p o on dono . This is in s a k con as wi h
he s uc u e o monosaccha ide-benzene/ oluene dime s, which
a e mainly o med by CHpin e ac ions. The eason o such a
diffe en beha iou is he modula ion in oduced by he pheno-
lic hyd oxyl g oup: he o ma ion o an in e molecula hyd ogen
bond seems o guide he whole agg ega ion p ocess, imposing
es ic ions in he con o ma ional landscape.
The o he de e minan ac o o he s uc u e adop ed in he
agg ega e is he posi ion o he O4H, axial in galac ose and
equa o ial in glucose. Such small diffe ence p opaga es
h ough he in e molecula hyd ogen bond ne wo k and de e -
mines he o ien a ion o he O6H g oup, o which phenol is
ancho ed, s ongly in luencing i s inal posi ion. This obse a-
ion is in ag eemen wi h p e ious s udies ha a ibu ed he
in amolecula hyd ogen bond ne wo k o a kind o ampli ica-
ion effec ha acili a es he ‘‘ eading’’ o he small s uc u al
diffe ences be ween suga s by he ecep o , imp o ing he
sensi i i y and speci ici y o he suga – ecep o in e ac ion.
Con lic s o in e es
The e a e no con lic s o decla e.
Fig. 5 S eng h o he hbonds in he sys ems s udied in his wo k s. shi
in he posi ion o he OH s e ching. The hbond s eng h was es ima ed
using he bond c i ical poin s. The numbe s co espond o he s uc u es in
Fig. 3. The alues calcula ed o he O4HO
Ph
H in e ac ion in s uc u es 7
and 8 we e added in ed. The ed ci cle labelled as ‘‘F ee Glc/Gal’’
co esponds o he alues o he OH g oups no di ec ly bonded o PhOH
in Glc and Gal. Values o PhOH and wa e OH s e ching we e also added
o compa ison.
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Acknowledgemen s
G an s PGC2018-098561 and PID2021-127918NB-I00 unded by
MCIN/AEI/10.13039/501100011033 and, by ‘‘ERDF A way o
making Eu ope’’. G an IT1491-22 unded by he Basque Go -
e nmen . F. T. H. acknowledges inancial suppo om Spanish
Minis y o Science and Inno a ion unde he FPI p edoc o al
p og am. A. C. would like o hank he Basque Go e nmen o
a p edoc o al ellowship. We also hank he SGIKER (UPV/EHU,
MICIU-FEDER) o he compu a ional and lase esou ces.
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