4. ENERGY-RICH COMPOUNDS
123
the primary energy source for many physiological processes. To a much
lesser extent, the other nucleoside triphosphates (NTP) may act as
phosphorylating agents for specific reactions, or they may transphosphorylate (51-54b) with ADP to yield ATP:
NTP + ADP ^ NDP + ATP
(23)
ATP was first isolated as the barium salt from muscle extracts in
1929 by Lohmann (55) and by Fiske and SubbaRow (56). The "energy-rich" nature of ATP was recognized by Meyerhof on the basis of
calorimetric measurements (31, 32) of the ΔΗ of hydrolysis and calculations (28) of the AF° of hydrolysis. ATP has been isolated from
muscle by a number of procedures (57-60), all employing fractionation with heavy metals and solvents. More recently, anion-exchange
chromatography (61) using gradient elution (62) has been used with
considerable success for both the isolation and the identification of ATP
and other nucleoside polyphosphates. ADP is customarily prepared by
the careful hydrolysis of ATP using an ATPase isolated from lobster
muscle (57, 63, 64). Adenosine tetraphosphate has been isolated (65,
66) from horse muscle and shown to be a linear 5'-tetraphosphate of
adenosine; as yet, no metabolic function has been ascribed to this
compound.
The chemical synthesis of ADP and ATP has been achieved by Todd
and his colleagues (67, 68) through the phosphorylation of the isopropylidine derivative of AMP with dibenzyl phosphorochloridate, followed by removal of the various protective groups. Improved methods,
employing dehydrating agents such as the carbodiimides (69, 69a),
were also utilized for the direct combination of phosphoric acid with
AMP or ADP. The chemical synthesis of UDP and UTP by the carbodiimide method has been reported (70).
ATP, ADP, adenosine tetraphosphate, and other nucleoside di- and
triphosphates may be separated from one another and quantitatively
estimated by means of ion-exchange chromatography (61, 62, 71). This
method is undoubtedly the most sensitive one available and is preferable for complex mixtures of these materials. Paper chromatography,
likewise, has been used in many investigations (see, for example, Bock
et al., 72) for the separation and identification of these nucleotides.
Colorimetric methods, based upon the release of Pi after 7-minute
hydrolysis at 100° in 1N acid, or based upon solubility in bariumalcohol fractionation procedures, are relatively nonspecific, and have
been largely replaced by the more sensitive Chromatographie methods.
Enzymatic assays (73-76), based upon the following groups of reac-
123
the primary energy source for many physiological processes. To a much
lesser extent, the other nucleoside triphosphates (NTP) may act as
phosphorylating agents for specific reactions, or they may transphosphorylate (51-54b) with ADP to yield ATP:
NTP + ADP ^ NDP + ATP
(23)
ATP was first isolated as the barium salt from muscle extracts in
1929 by Lohmann (55) and by Fiske and SubbaRow (56). The "energy-rich" nature of ATP was recognized by Meyerhof on the basis of
calorimetric measurements (31, 32) of the ΔΗ of hydrolysis and calculations (28) of the AF° of hydrolysis. ATP has been isolated from
muscle by a number of procedures (57-60), all employing fractionation with heavy metals and solvents. More recently, anion-exchange
chromatography (61) using gradient elution (62) has been used with
considerable success for both the isolation and the identification of ATP
and other nucleoside polyphosphates. ADP is customarily prepared by
the careful hydrolysis of ATP using an ATPase isolated from lobster
muscle (57, 63, 64). Adenosine tetraphosphate has been isolated (65,
66) from horse muscle and shown to be a linear 5'-tetraphosphate of
adenosine; as yet, no metabolic function has been ascribed to this
compound.
The chemical synthesis of ADP and ATP has been achieved by Todd
and his colleagues (67, 68) through the phosphorylation of the isopropylidine derivative of AMP with dibenzyl phosphorochloridate, followed by removal of the various protective groups. Improved methods,
employing dehydrating agents such as the carbodiimides (69, 69a),
were also utilized for the direct combination of phosphoric acid with
AMP or ADP. The chemical synthesis of UDP and UTP by the carbodiimide method has been reported (70).
ATP, ADP, adenosine tetraphosphate, and other nucleoside di- and
triphosphates may be separated from one another and quantitatively
estimated by means of ion-exchange chromatography (61, 62, 71). This
method is undoubtedly the most sensitive one available and is preferable for complex mixtures of these materials. Paper chromatography,
likewise, has been used in many investigations (see, for example, Bock
et al., 72) for the separation and identification of these nucleotides.
Colorimetric methods, based upon the release of Pi after 7-minute
hydrolysis at 100° in 1N acid, or based upon solubility in bariumalcohol fractionation procedures, are relatively nonspecific, and have
been largely replaced by the more sensitive Chromatographie methods.
Enzymatic assays (73-76), based upon the following groups of reac-