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Metabolism — glycolysis (step 1)

Hexokinase: glucose phosphorylation

How the Hexokinase works, in 3 steps. Each step shows the structures and the curved arrows that move the electrons.

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In short. A 3-step mechanism. Typical reagents: Glucose, Mg·ATP, hexokinase. Conditions: Aqueous, pH ~7.4, 37 °C. Bonds formed: C6–O–P ester; ADP; bonds broken: O–H (C6); P–O (γ-P to bridging O). Stereochemistry: In-line SN2-like, inversion at P. Rate and selectivity: Irreversible (ΔG°′ ≈ −17 kJ/mol); glucose induces domain closure.

At a glance

ReagentsGlucose, Mg·ATP, hexokinase
Solvent / conditionsAqueous, pH ~7.4, 37 °C
Bonds formedC6–O–P ester; ADP
Bonds brokenO–H (C6); P–O (γ-P to bridging O)
StereochemistryIn-line SN2-like, inversion at P
Rate / selectivityIrreversible (ΔG°′ ≈ −17 kJ/mol); glucose induces domain closure

Step 1: Base deprotonates the C6–OH

+EnzOOHOOOHOHOHOH1 · BASE DEPROTONATES THE C6–OHAsp carboxylateBASEglucoseC₆H₁₂O₆SUBSTRATEInduced fit positions C6–OH next to the base and ATP.

Glucose's C6 hydroxyl, positioned by the enzyme's induced-fit closure around the substrate, is deprotonated by a nearby active-site base, activating it as a nucleophile.

Step 2: In-line attack on the γ-phosphate

+OOOHOHOHOHADPOPOOO2 · IN-LINE ATTACK ON THE Γ-PHOSPHATEglucose 6-alkoxideC₆H₁₁O₆⁻NUCLEOPHILEMg·ATP(γ-phosphate shown)ELECTROPHILEattackin-line: oppositethe ADP leaving groupPentacoordinate phosphorane-like TS; ADP leaves on the opposite side.

That activated oxygen attacks ATP's terminal (γ) phosphorus in-line, opposite the ADP leaving group — the identical pentacoordinate-phosphorane phosphoryl-transfer mechanism covered generally much earlier in this library, here run by the very first committed enzyme of glycolysis.

Step 3: Glucose 6-phosphate + ADP

+OPOOOOOHOHOHOHAdoOPOOOPOOO3 · GLUCOSE 6-PHOSPHATE + ADPglucose 6-phosphateC₆H₁₁O₉P²⁻PRODUCTADPBYPRODUCTCharged G6P cannot cross the membrane: it is trapped and committed to glycolysis.

The products are glucose-6-phosphate and ADP — this irreversible, energy-investing step traps glucose inside the cell (phosphorylated sugars can't cross the membrane) and commits it to the glycolytic pathway.

Abbreviations and terms

TS
Transition state. The highest-energy point on the pathway between two minima; it cannot be isolated.
SN2
Bimolecular nucleophilic substitution. One concerted step with backside attack and inversion; rate = k[RX][Nu].
ATP
Adenosine triphosphate. The cell’s energy currency; transfers phosphate groups.
ADP
Adenosine diphosphate. ATP after losing one phosphate.
Asp
Aspartate. Acidic amino acid.

How it compares with related reactions

HexokinaseEnolaseGAPDH
ReagentsGlucose, Mg·ATP, hexokinase2-Phosphoglycerate, 2 Mg²⁺, enolase (Lys345 base, Glu211 acid)Glyceraldehyde-3-phosphate (G3P), NAD⁺, inorganic phosphate (Pi), GAPDH (Cys149, His176)
ConditionsAqueous, pH ~7.4, 37 °CCytosol, pH ≈ 7, 37 °CAqueous, pH ≈ 7.4, 37 °C (cytosol)
StereochemistryIn-line SN2-like, inversion at PAnti elimination of H and OH; reversibleHydride delivered to one face of NAD⁺ (pro-R)
Rate / selectivityIrreversible (ΔG°′ ≈ −17 kJ/mol); glucose induces domain closureΔG°′ ≈ +1.8 kJ/mol; PEP has a very high phosphoryl-transfer potentialFast and reversible; couples oxidation to phosphate capture

Test yourself

Try to answer each question before opening it.

Which reagents and conditions are typical for the Hexokinase?

Glucose, Mg·ATP, hexokinase. Conditions: Aqueous, pH ~7.4, 37 °C.

Which bonds are formed and which are broken?

Formed: C6–O–P ester; ADP. Broken: O–H (C6); P–O (γ-P to bridging O).

What is the stereochemical outcome?

In-line SN2-like, inversion at P

What controls the rate or the selectivity?

Irreversible (ΔG°′ ≈ −17 kJ/mol); glucose induces domain closure

Study guides that help with this reaction

Related mechanisms

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