An ab initio molecular dynamics investigation

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HOW DOES NATURE FORM HOW DOES NATURE FORM

GLYCOSIDIC BONDS?

An ab initio molecular dynamics investigation

C R i

An ab initio molecular dynamics investigation

Carme Rovira

Universitat de Barcelona – Parc Científic de Barcelona Parc Científic de Barcelona

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Carbohydrates

50% of our daily

calorie intake comes from carbohydrates

http://en wikipedia org/

Carbohydrates are our “biological fuel “,

http://en.wikipedia.org/

as well as the primary form of storage and energy consumption in organisms

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Introduction

The roles of carbohydrates

P l h id ^starch

Polysaccharides

 Structural support

E t

starch cellulose

Glycoconjugates

 Energy storage

Glycoconjugates

 Cell-cell interaction

 Signal transduction

 Immune response

 Parasitic infections carbohydrates

 Parasitic infections

GlcNAcMan₅GlcNAc₂

carbohydrates http://www.glycomicscentre.ca

D i h i h i i hi h b h d i li d

Deciphering mechanisms in which carbohydrates are implicated is of enormous interest for the search of new therapeutic agents.

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Introduction

Glicosidic bond

O OH

OH

OH O

O HO

O O

HO OH

OH OH

O HO

O O

glucose glucose glucose glucose

HO O

HO

glycosidic bond:

C O bond between two sugar units C-O bond between two sugar units

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How do glycosidic bonds form?

Most glycosidic bonds are synthesized in nature from sugars that are activated by a cofactor

Enzyme (glycoside transferase)

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How do glycosidic bonds form?

The glycosidic bond is formed upon transfer of a sugar molecule from g y p g the donor (an activated sugar) to an acceptor molecule (typically another sugar)

Enzyme (glycoside transferase)

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Two modes of enzyme operation

Retention or inversion of the configuration of the anomeric carbon

t i i GT retaining GT

The molecular mechanism of retaining GTs is very controversial

Palcic, Curr. Opin. Chem. Biol. 2011; Lee et al. Nat. Chem. Biol. 2011 Lairson et al. Annu. Rev. Biochem. 2008

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Retention of the configuration of the anomeric carbon

t i i GT retaining GT

high steric hindrance is expected

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Possible mechanism for retaining enzymes

covalent glycosyl-enzyme intermediate

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Possible mechanism for retaining enzymes

covalent glycosyl-enzyme intermediate (double displacement,

~ retaining GHs)

retaining GHs

e.g. Biarnés et al. J. Am. Chem.

Soc. 133, 20301–09, 2011

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Possible mechanism for retaining enzymes

covalent glycosyl-enzyme intermediate (~ retaining GHs)

But

• All experimental attempts to isolate a glycosyl-enzyme intermediate have failed

• Few GTs have a putative

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Another possibility

+ -

The reaction takes place on a single “face” of the sugar

“front-face attack”

g g

B t

• High steric hindrance expected

But

• Little chemical precedence

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Controversy

Two covalent bonds being broken/formed in the

+ -

broken/formed in the

same region of the space

I th f t f ti f ibl ?

• Is the front-face reaction feasible?

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Simulation model

•Ab initio molecular dynamics (to take into account the atomic

and electronic motion at room temperature) QM

p )

• QM/MM

(Density Functional Theory/ AMBER) MM (Density Functional Theory/ AMBER)

• Metadynamics (Laio and Parrinello, PNAS 99, 12562-66, 2002) (to model the chemical reaction)

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Enzyme studied: trehalose-6-phosphate synthase

O OH

OPO^2-

O HO

HO

OH

O OH OH OH

O O

O P

O^- O

O

+

O OH OH

N

P O^- O

O

+

HO HO

HO

O HO

OH OH

OPO₃

OH

HO HO

O HO

OH O OPO₃^2-

OH

P O^- O

O O

N

HN O

O

P O^- O

enzyme O

O

UDP-glucose (donor)

glucose-6P (acceptor)

N

HN O

O

O O

trehalose-6P UDP

O

Enzymey Trehalose is a natural disaccharideTrehalose is a natural disaccharide used as food ingredient for its

sweet flavor and preservative properties

properties

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Enzyme·substrate complex

St t t bl d l l d i

• Structure stable under molecular dynamics

• Good agreement with binary complexes structures

(Enzyme + UDP-Glc and Enzyme + UDP + Glc-6P) (Enzyme + UDP-Glc and Enzyme + UDP + Glc-6P)

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Free energy landscape

metastable intermediate

~ 100 QM atoms

20 ps AIMD, 10⁵ h MN (64/128 procs).

lifetime ~ 2 ps

(dos milésimas de una milmillonésima de segundo!)

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R

cleavage of phosphate-sugar bond

glycosidic bond

formation

1 1

2

3 R 3

formation

4

P

P P

proton transfer

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Molecular mechanism of the front-face reaction

metadynamics metadynamics

trajectory

Glucose-6P

UDP UDP

Theory: Ardèvol & Rovira, Angew. Chem. Int. Ed. 50, 10897 –901, 2011 Experiment: Seung et al. Nat. Chem. Biol. 7, 631-38, 2011

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Acknowledgments

Albert Ardèvol (ETH, Switzerland)

Discussions with:

Antoni Planas (Universitat Ramon Llull Barcelona) Antoni Planas (Universitat Ramon Llull, Barcelona) Seung Lee, Ben Davis (University of Oxford, UK)