kinase it is a commonly used regeneration system [565]. However, acetyl
phosphate is modestly stable in aqueous solution (t 1/2 at pH 4.2–7.2
ca. 7–21 h) and its phosphoryl donor potential is lower than that of PEP. As
for pyruvate kinase, acetate kinase also can accept nucleoside phosphates other
than adenosine, and it is inhibited by acetate. Regeneration of other nucleoside
triphosphates (GTP, UTP, and CTP) or the corresponding 2
0 -deoxynucleoside
triphosphates – which are important substrates for enzyme-catalyzed glycosyl
transfer reactions (Sect. 2.6.1) [561, 566, 567] – can be accomplished in the
same manner using the PEP- or acetate kinase systems.
• As an alternative to expensive phosphenol pyruvate and hydrolytically unstable
acetyl phosphate, the use of creatine kinase together with creatine phosphate has
been proposed for ATP-recycling [568]. Due to the (controversially debated) role
of creatine as food supplement for bodybuilding, creatine phosphate is relatively
inexpensive and it is an equally strong phosphate donor as acetyl phosphate.
• Promising ATP-recycling methods for large-scale applications use cheap inorganic polyphosphate as phosphate donor and polyphosphate kinase, respectively
[569, 570]. Polyphosphate is a ubiquitous natural polymer of tens to hundreds of
orthophosphate residues linked by a high-energy phosphoanhydride bond, which
is believed to be an ancient energy carrier preceding ATP in the prebiotic age
[571]. It is widely used as an acidulant additive in soft drinks. Polyphosphate
kinase from E. coli accepts also other nucleoside diphosphates and yields up to
40 regeneration cycles [572], but owing to the limited phosphate donor strength
of poly/pyrophosphate, equilibrium yields for ATP are !85% [573].
OH
O
O
OH
O
OH
Substrate
OH
Donor
O
Donor
O
Substrate
CO 2 H
O
O
O
ADP
ATP
regenerating
kinase
phosphorylating
kinase
acetate kinase
pyruvate kinase
Donor-OH
Donor-Phosphate
polyphosphate kinase
n
n = 11-16
P
P
P
P
P
P
P
P
P
NH
N
H
P
N
HO 2 C
NH
NH 2
N
HO 2 C
creatine kinase
n
Scheme 2.75 Use of kinases for the enzymatic phosphorylation of alcohols and ATP recycling
108
2 Biocatalytic Applications
phosphate is modestly stable in aqueous solution (t 1/2 at pH 4.2–7.2
ca. 7–21 h) and its phosphoryl donor potential is lower than that of PEP. As
for pyruvate kinase, acetate kinase also can accept nucleoside phosphates other
than adenosine, and it is inhibited by acetate. Regeneration of other nucleoside
triphosphates (GTP, UTP, and CTP) or the corresponding 2
0 -deoxynucleoside
triphosphates – which are important substrates for enzyme-catalyzed glycosyl
transfer reactions (Sect. 2.6.1) [561, 566, 567] – can be accomplished in the
same manner using the PEP- or acetate kinase systems.
• As an alternative to expensive phosphenol pyruvate and hydrolytically unstable
acetyl phosphate, the use of creatine kinase together with creatine phosphate has
been proposed for ATP-recycling [568]. Due to the (controversially debated) role
of creatine as food supplement for bodybuilding, creatine phosphate is relatively
inexpensive and it is an equally strong phosphate donor as acetyl phosphate.
• Promising ATP-recycling methods for large-scale applications use cheap inorganic polyphosphate as phosphate donor and polyphosphate kinase, respectively
[569, 570]. Polyphosphate is a ubiquitous natural polymer of tens to hundreds of
orthophosphate residues linked by a high-energy phosphoanhydride bond, which
is believed to be an ancient energy carrier preceding ATP in the prebiotic age
[571]. It is widely used as an acidulant additive in soft drinks. Polyphosphate
kinase from E. coli accepts also other nucleoside diphosphates and yields up to
40 regeneration cycles [572], but owing to the limited phosphate donor strength
of poly/pyrophosphate, equilibrium yields for ATP are !85% [573].
OH
O
O
OH
O
OH
Substrate
OH
Donor
O
Donor
O
Substrate
CO 2 H
O
O
O
ADP
ATP
regenerating
kinase
phosphorylating
kinase
acetate kinase
pyruvate kinase
Donor-OH
Donor-Phosphate
polyphosphate kinase
n
n = 11-16
P
P
P
P
P
P
P
P
P
NH
N
H
P
N
HO 2 C
NH
NH 2
N
HO 2 C
creatine kinase
n
Scheme 2.75 Use of kinases for the enzymatic phosphorylation of alcohols and ATP recycling
108
2 Biocatalytic Applications