the enzymatic acetylation of amines the enzymatic

THE ENZYMATIC ACETYLATION OF AMINES

BY HERBERT TABOR, ALAN H. MEHLER, AND E. R. STADTMAN

(From the National Znstitute of Arthritis and Metabolic Diseases and the National Heart Znstitute, National Institutes of Health, United States Public

Health Service, Bethesda, Maryland)

(Received for publication, February 16, 1953)

The enzymatic acetylation of aromatic amines (1, Z), choline (3), glucosamine (4), and histamine (5) has been shown to require coenzyme A (CoA) ; recent studies (6) have demonstrated that the thio ester, acetyl CoA, can serve as the acetyl donor in the acetylation reaction. However, more detailed studies on the properties of the acetylating enzyme were made difficult by the absence of a suitable method for following the acetylation reaction continuously. In the work reported here, a convenient, direct spectrophotometric method was used for following the acetylating reaction, which depended on the marked difference between the absorption spectra of free and acylated aromatic amines previously used in the study of the hydrolysis of formylkynurenine (7). Most of the studies were carried out with p-nitroaniline, as the changes in absorption produced by acetylation could be measured above 400 mM; this wave-length is sufficiently high so that interference from the extraneous absorption commonly found in tissue extracts is largely avoided. With purified enzyme preparations the spectrophotometric method can also be used with a variety of other compounds. Using the spectrophotometric method, we have studied acetylation of amines by acetyl CoA, the sulfhydryl nature of the acetylating enzyme, and inhibition of the acetylation reaction by free CoA.

Methods and Materials

CoA, containing approximately 1 PM per mg. (approximately 320 units per mg. (8)), was obtained from the Pabst Brewing Company. Acetyl CoA was prepared by treating reduced CoA with thioacetic acid (9); the reaction product was purified by paper chromatography (10). Butyryl CoA was similarly prepared with thiobutyric acid. We are indebted to Dr. A. Kornberg for a sample of palmityl CoA, to Dr. H. Bauer for acetyl- p-aminobenzoic acid, to Dr. W. Kielley for p-chloromercuribenzoate, and to Dr. 0. Hayaishi for kynurenine. Acetylhistamine (ll), acetyl phosphate (X2), and transacetylase (13) were prepared as previously described. Samples of disodium ethylenediaminetetraacetate (analytical grade) (EDTA), the various amines, and diphosphopyridine nucleotide (DPN) were obtained commercially. p-Nitroaniline, p-nitroacetanilide, and m-ni-

127

by guest on April 3, 2018

http://ww

w.jbc.org/

Dow

nloaded from

http://www.jbc.org/

128 ENZYMATIC ACETYLATION OF AMINES

troaniline were recrystallized prior to use. Crystalline condensing enzyme (14) was prepared by Dr. J. Stern. Malic dehydrogenase was purified approximately 45-fold from Pseudomonas jluorescens extracts by ammonium sulfate and alumina CT fractionations.

All spectrophotometric measurements were carried out in a Beckman DU spectrophotometer in cuvettes with a 1 cm. light path at 25. The curves representing direct experiments have been corrected for the changes in optical density caused by dilution upon the addition of various reagents, Protein concentrations were usually measured spectrophotometrically at

E I? k :

1.0

0

$ 5 z 0.5 F w

%

0 0 2 260 340 420 500 220 260 300 340

WAVE LENGTH mp WAVE LENGTH mp

FIG. 1. Absorption spectra of p-nitroaniline, p-nitroacetanilide, p-aminobenzoic acid, and acetyl-p-aminobenzoic acid. A, p-nitroaniline (0) and p-nitroacctanilide (0); R, p-aminobenzoic acid (0) and acetyl-p-aminobenzoic acid (0). (The optical densities were measured at a concentration of 8 X 10e5 M in 0.04 M potassium phosphate buffer, pH 6.8.)

280 rng; the optical density at 260 rnp was used to correct for nucleic acid (15). Ninhydrin determinations were carried out by the reduced ninhydrin method of Moore and Stein (16).

EXPERIMENTAL

The absorption spectra of p-aminobenzoic acid (PABA) and p-nitroaniline (PNA) are shown in Fig. 1, together with the spectra of the N-acetyl derivatives. When PABA was used, the acetylation reaction was followed at 300 rnp, which was the lowest wave-length at which absorption of the enzymes did not interfere excessively. With PNA, it was usually found convenient to measure the changes in absorption at 420 mp, in order to permit substrate concentrations of 10e4 M to be used with


http://ww

w.jbc.org/

Dow

nloaded from

http://www.jbc.org/

H. TABOR, A. H. MEHLER, AND E. R. STADTMAN 129

initial density readings of 0.6, although larger differences in absorption can be obtained at other wave-lengths.

Assay of Acetylating Enzyme-The enzyme was assayed in a volume of 1 ml. at pH 6.8 and an initial concentration of 1O-4 M PNA. A unit of enzyme is defined as the amount of enzyme causing a decrease of 0.001 unit of optical density per minute at 420 mp. The proportionality of the rate of change in optical density to enzyme concentration is shown in Fig. 2. Specific activity is defined as units per mg. of protein. The effects of varying the concentration of CoA and PNA are shown in Fig. 3.

MINUTES

FIG. 2. Acetylation of p-nitroaniline with varying amounts of crude pigeon liver extracts. Incubation mixture contained 40 JJM of potassium phosphate buffer (pH 6.8), 5 PM of sodium thioglycolate, 12 pM of dilithium acetyl phosphite, 4 units of transacetylase, 0.12 pM of CoA, 0.1 PM of p-nitroaniline, crude pigeon liver extract, and water to a total volume of 1.05 ml. The numbers on the curves represent ml. of enzyme used. The optical density of 0.1 pM of p-nitroaniline in 1.05 ml. is 0.571.

Since less than saturating concentrations of PNA were used routinely, only initial rates were used in enzyme assays, and levels of enzyme were selected to allow rates to be determined before the substrate concentration changed by more than 10 per cent.

Purification of Acetylating Enzyme-An acetone powder of pigeon liver was prepared by blending freshly removed liver twice with 10 volumes of cold (- 10) acetone each time, filtering with suction, and air drying at room temperature. The powders were stored in the refrigerator.

12 gm. of the liver powder were extracted with 120 ml. of distilled water at room temperature by grinding in a mortar for 10 to 15 minutes. After centrifugation at 22,000 X g for 10 minutes, the supernatant fluid was cooled to 0. This extract was immediately fractionated with acetone


http://ww

w.jbc.org/

Dow

nloaded from

http://www.jbc.org/


and alumina Cy (17). Acetone had previously been used in the fractiona- tion of this enzyme (18).

To 96 ml. of extract (39,000 units; specific activity lO.S), 76 ml. of cold acetone were added (0). After centrifugation, the precipitate was dis- carded and 193 ml. of acetone were added to the supernatant fluid. The precipitate was collected by centrifugation and dissolved in 15 ml. of cold water (26,000 units). The enzyme was then adsorbed on 90 ml. of alumina gel CT (11 mg. of solids per ml.), which was centrifuged and washed with 100 ml. of water. The enzyme was eluted with 100 ml. of 0.01 M

MICROMOLES OF CoA MICROMOLES OF p-NITROANILINE

FIG. 3. Affinity of the acetylating enzyme for CoA and p-nitroaniline. The incubation mixtures contained 40 PM of potassium phosphate (pH 6.8), 5 PM of sodium thioglycolate, 13 PM of dilithium acetyl phosphate, and 4 units of transacetylase. With varying concentrations of CoA (Fig. 3, A) 0.1 PM of PNA and 10.3 units of acetylating enzyme were included. In the case of varying PNA concentration (Fig. 3, B), 5 pM of EDTA, 0.06 PM of CoA, and 23 units of acetylating enzyme were added. The final volume was 1.0 ml. Initial rates were determined at 405, 420, or 430 rnp, and are expressed in micromoles per minute.

potassium phosphate buffer (pH 7.8) in three portions (15,000 units; specific activity 275). Additional activity was obtained by further elu- tion with 50 ml. of the buffer (3500 units; specific activity 184).

Enzyme solutions at this stage of purification were used in all experiments, unless otherwise indicated. The activity of the frozen enzyme solutions was essentially unchanged at - 15 for at least 3 months. More concentrated enzyme solutions could be obtained by lyophilization after the addition of 100 PM of EDTA per 100 ml.

Acetyl CoA-When excess acetyl phosphate was used as acetyl donor, the reaction proceeded until no free amine was detected, and the spectrum of the corresponding p-nitroacetanilide was obtained. In Fig. 4 it is shown that the reaction also goes to completion with respect to acetyl


http://ww

w.jbc.org/

Dow

nloaded from

http://www.jbc.org/


CoA, when the amine is present in higher concentration. In the presence of cysteine or thioglycolate in slightly alkaline solution (pH S), however, the reaction ceases before the theoretical value has been attained. This can be explained by the loss of acetyl CoA by the non-enzymatic acetylation of sulfhydryl compounds previously described (19).

A confirmation of the nature of the reaction was obtained by measuring the disappearance of acetyl CoA during the acetylation reaction. This was done by determining the acetyl CoA at various times with the citrate- condensing enzyme which forms citrate from oxalacetate and acetyl CoA

0 20 40 60 80 100 MINUTES

FIG. 4. Stoichiometric reaction between added acetyl CoA and p-nitroaniline. Incubation mixture contained 100 PM of potassium phosphate (pH 6.8), 5 pM of sodium thioglycolate, 0.1 pM of p-nitroaniline, 0.0445 j.&M of acetyl CoA, and 25 units of acetylating enzyme in a total volume of 1.0 ml. At the arrow, an additional 0.022 PM of acetyl CoA (0.05 ml.) was added to the incubation mixture. The dotted lines represent the theoretical decrease in optical density for each acetyl CoA addition.

(13). Stern et al. (20) have reported studies on the coupling of the malic dehydrogenase and citrate condensation systems. In the combined system, malate is oxidized by DPN to form oxalacetate and DPNH. The oxalacetate is then used to form citrate with acetyl CoA. The reduction of DPN in this system does not proceed to a measurable extent unless the oxalacetate is removed. From the equilibrium constants calculated by Stern et al., one can calculate that the formation of citrate from acetyl CoA in the presence of excess malate and DPN should proceed essentially to completion. Therefore, DPN reduction should be a measure of acetyl CoA added. This was found to be the case; DPN reduction in the combined system, measured by the increase in density at 340 rnp (21), agreed with the acetyl CoA added, as determined by the arsenolysis reaction with


http://ww

w.jbc.org/

Dow

nloaded from

http://www.jbc.org/


phosphotransacetylase (22). The validity of this method for following acetylations was shown in an experiment with PABA. 0.185 PM of acetyl CoA was incubated with acetylating enzyme, substrate, and buffer until 0.077 pM of PABA had been acetylated, as measured by the change in optical density at 300 mp. An aliquot of this mixture was added to a vessel containing 8 pM of dl-malic acid, 0.2 PM of DPN, 6000 units of malic dehydrogenase, and 20 pM of phosphate buffer, pH 6.8, in a total volume of 2.9 ml. The amount of acetyl CoA present was measured by the reduction of DPN after the addition of 0.1 ml. of condensing enzyme. The results indicated that 0.087 pM of acetyl CoA had disappeared during the prior reaction with PNA.

When butyryl CoA was substituted for acetyl CoA in an incubation mixture similar to that used in Fig. 4, the rate of acylation was only 4 per cent of that observed with acetyl CoA. When palmityl CoA was added as the sole acylating agent, no acylation was observed. Furthermore, addition of 0.1 pM of palmityl CoA to an incubation mixture containing 0.2 PM of acetyl CoA resulted in complete inhibition of the acetylating reaction; 0.01 PM of palmityl CoA produced approximately 50 per cent inhibition.

As shown in Fig. 5, the presence of free CoA inhibits the acetylation reaction with acetyl CoA. These results were obtained with the com- mercial preparation. Essentially similar findings were found with an- other CoA preparation which had been prepared by a different procedure involving ion exchange chromatography on Dowex 1 and 50 (10).

pH Dependence-From pH values below 6 to over 9.5, the activity is essentially constant when p-nitroaniline is used as the substrate (Fig. 6). With histamine, a stronger base, the reaction falls off sharply below pH 8.5. Although this suggests that only the uncharged form of the amino group can be acetylated, a definite statement cannot be made, without knowing whether the same enzyme carried out both acetylations. It is of interest that Koshland (23) reported a pH dependence for the non-enzymatic acetylation of glycine by acetyl phosphate. At the concentrations of substrate and acetyl phosphate used in our experiments, the non- enzymatic reaction did not introduce a significant error.

Sulfhydryl Requirement-In slightly alkaline incubation mixtures, an absolute requirement for a reducing agent, such as sodium thioglycolate, cysteine, or hydrogen sulfide, can be demonstrated (Fig. 7, A). This requirement was not found if the enzyme had been preincubated with EDTA (Fig. 7, B), and thus is presumably associated with a metal impurity in the incubation mixture. EDTA can only partially reactivate an inactivated enzyme, but essentially complete reactivation can be ac- complished by the addition of reducing agent. A less pronounced effect


http://ww

w.jbc.org/

Dow

nloaded from

http://www.jbc.org/

HI. TABOR, A. H. MEHLER, AND E. R. STADTMhN 133

of reducing agents is seen at pH 6.8 (Fig. 7, C), at which there is essentially no effect on the initial rate of the reaction, even after preincubation of the entire reaction mixture for 28 minutes at 25. This is consistent with the well known effect of pH on sulfhydryl oxidations (24). In some experiments reducing agent alone was not sufficient to maintain the enzyme activity; EDTA was required in addition.

0 IO 20 30 40 50 .OlI I I I I I

5 6 7 8 9 IO MINUTES PH

FIG. 5 FIG. 6

FIG. 5. Inhibition by free CoA of acetylation by acetyl CoA. The incubation mixture contained 400 pM of potassium phosphate buffer, pH 6.8, 5 FM of sodium thioglycolate, 5 pM of EDTA, 0.045 PM of acetyl CoA, 0.1 pM of p-nitroaniline, free CoA as indicated by the numbers on the curves (in micromoles), and 7.5 units of acetylating enzyme in a final volume of 1 .O ml.

FIG. 6. Effect of pH on acetylation of p-nitroaniline. The incubation mixture contained 5 pM of EDTA, 5 pM of sodium thioglycolate, 40 units of acetylating enzyme, 0.09 PM of acetyl CoA, 0.1 PM of p-nitroaniline, and 20 to 400 PM of the appro- priate buffer in a final volume of 1.0 ml. The initial velocities are expressed as the

decrease in optical density per minute at 420 m+ 0, potassium acetate; @, potassium phosphate; 0, tris(hydroxymethyl)aminomethane (Tris); A, sodium pyrophosphate; V, potassium glycine; 0, potassium borate.

The sulfhydryl nature of the acetylating enzyme is further shown in Fig. 8, in which complete inhibition was caused by a final concentration of 1O-5 M p-chloromercuribenzoate (25). This inhibition was completely reversed by sodium thioglycolate.

Other Substrates--With essentially the same conditions as those used for p-nitroaniline acetylation (1 ml. volume, containing 0.1 pM of substrate, 13 PM of acetyl phosphate, transacetylase, and 0.1 pM of CoA), spectro-

1 Independently, S. P. Bessman (personal communication), working in the laboratory of F. Lipmann, has also noted the effect of sulfhydryl compounds and EDTA on the transfer of acetyl groups of CoA by purified acceptor enzymes. 4-Aminoazo- benzene-4-sulfonic acid is employed as an acetyl acceptor.


http://ww

w.jbc.org/

Dow

nloaded from

http://www.jbc.org/

134 ENZYhlATIC! ACETYLATION OF AMINES

photometric methods were employed to follow the acetylation of a number of other aromatic amines. Under these conditions, the initial reaction

MINUTES MINUTES MINUTES

FIG. 7. Effects of EDTA and sulfhydryl compounds on the acetylating enzyme. A, effect of sodium thioglycolate on the acetylstion of p-nitroaniline at pH 8. The incubation mixtures contained 200 PM of Tris buffer, 0.1 MM of p-nitroaniline, 8 units of acetylating enzyme (in 0.2 ml. containing 20 PM of potassium phosphate) in a total volume of 0.9 ml. (pH 8). The incubation mixture of Experiment I contained, in addition, 5 PM of sodium thioglycolate. At arrow a, 0.044 PM of acetyl CoA (0.1 ml.) was added to each incubation mixture. At arrow b, 5 pM of sodium thioglycolate (0.05 ml.) were added to the incubation mixture of Experiment II. The theoretical decrease in optical density for 0.044 PN in 1 ml. is 0.264. B, effect of EDTA on the acetylation of p-nitroaniline at pH 8. The incubation mixture contained 200 PM of Tris buffer, pH 8.1, 0.1 pM of p-nitroaniline, 16 units of acetylating enzyme (in 0.1 ml. containing 6pM of phosphate) in a total volume of 0.9 ml. (pH 8). 10 FM of EDTA were also included in the incubation mixture for Experiment I. After 20 minutes incubation at 25 0.044 I.~M of acetyl CoA (0.1 ml.) was added to both Experiments I and II (at arrow a). At arrow b, 10 PX of EDTA (0.1 ml.) were added to Experiment II; at arrow c, 5 PM of cysteine (0.05 ml.) were added to Experiment II. C, effect of sodium thioglycolate on the acetylation of p-nitroaniline at pH 6.8. The incubation mixture contained 25 PM of potassium phosphate buffer (pH 6.8), 0.1 PM of p-nitroaniline, and 7 units of acetylating enzyme in a total volume of 0.9 ml. In Experi- ment I, 5 PM of sodium thioglycolate were also present. At arrow a, 0.044 PM of acetyl CoA (0.1 ml.) was added to each incubation mixture. The theoretical decrease in optical density is 0.264; the final decrease attained with the incubation mixture of Experiment I was 0.253.

rates with p-aminobenzoic acid, m-nitroaniline, o-phenylenediamine, o-toluidine, m-toluidine, o-bromoaniline, p-bromoaniline, o-anisidine, and p- anisidine were found to be comparable to that with p-nitroaniline. On the other hand, no spectral changes were observed with o-aminobenzoic


http://ww

w.jbc.org/

Dow

nloaded from

http://www.jbc.org/


acid (anthranilic acid), o-nitroaniline, orthanilic acid, sulfanilic acid, p-nitromethylaniline, or kynurenine. p-Nitroaniline or p-anisidine added to these inactive incubation mixtures was acetylated, indicating that no pronounced inhibitory activity was present. It has been found convenient to use PNA at a concentration of low4 M, which is far below saturation, and the other substrates were studied at the same level. Since these substrates were not tested at the same relative concentrations with respect to enzyme saturation, further detailed comparisons of the actual rates obtained have not been presented.

da 0 E .05 F-0

%: z k 0.1 iti 4 lx

i

0.15

20 60 100 140 180 220 260

MINUTES FIG. 8. Inhibition of p-nitroaniline acetylation by p-chloromercuribenzoate. The

incubation mixture contained 40 pM of potassium phosphate buffer (pH 6.8), 5 PM of EDTA, 3 units of acetylating enzyme, and water to a total volume of 0.8 ml. The incubation mixture of Experiments II and III also contained p-chloromercuribenzoate (final concentration 1O-6 and IO-6 M, respectively). After 15 minutes incubation at 25, 0.1 pM (0.1 ml.) of p-nitroaniline and 0.09 PM (0.1 ml.) of acetyl CoA were added to the three incubation mixtures. At 147 minutes 5 PM of sodium thioglycolate were added to the incubation mixture of Experiment III.

The acetylation of histamine was followed by the disappearance of the ninhydrin-reacting material according to the method of Moore and Stein (16). Acetylhistamine does not react in this test. In incubation mixtures containing 2 PM of histamine, 300 PM of potassium pyrophosphate buffer, pH 9.3, 4 units of transacetylase, 10 pM of lithium acetyl phosphate, 0.1 PM of CoA, 10 pM of sodium thioglycolate, 5 pM of EDTA, and 160 units of acetylating enzyme in a total volume of 1.0 ml., approximately 50 per cent of the histamine is acetylated in 30 minutes. Essentially comparal)le data were obtained in other experiments, in which acetylhistamine was determined by a diazotization procedure (26), after differential extraction into tertiary butanol at pH 8 (27). Under similar conditions, 0.2 pM of phenylethylamine was acetylated in 30 minutes, as determined by the ninhydrin procedure.


http://ww

w.jbc.org/

Dow

nloaded from

http://www.jbc.org/


Previous studies on amine acetylations have used sulfanilamide (I), p-aminobenzoic acid (l), aminoazobenzene (28), and glucosamine (4). The purified enzyme preparations also acetylate glucosamine, but only at about 4 per cent of the rate of PNA acetylation. The report of Chou and Soodak (4) that this acetylation is not inhibited by benzylpenicillin has been confirmed, as well as the inhibition by penicillin of aromatic amine acetylation at low CoA concentrations reported by Soodak (29). In these experiments acetylglucosamine was determined by a micromodifi- cation of the Morgan and Elson method (30) devised by Dr. J. Strominger (personal communication).

DISCUSSION

The partially purified enzyme preparation described in this paper is capable of catalyzing the acylation of various amines by acetyl CoA and, more slowly, by butyryl CoA. The data presented show that, under certain conditions, the sulfhydryl nature of the enzyme can be demonstrated. The effects of thioglycolate and EDTA indicate that inactivation is probably associated with the sulfhydryl-binding and sulfhydryl- oxidizing properties of a heavy metal impurity. Although the r8les of essential sulfhydryl groups in the reaction mechanisms of many different kinds of enzymes are not known, it is tempting to consider that the SH group of the acetylating enzyme is involved in the formation of an acyl- enzyme as an intermediate in the transfer of acyl groups from CoA to amines, analogous to the mechanism suggested by Racker and Krimsky for triosephosphate dehydrogenase (31).

The apparent discrepancy between the results reported here on the requirement for sulfhydryl groups and the previous reports from other laboratories (6) of stoichiometric acetylation by acetyl CoA in the absence of added reducing agent may be explained by the following considerations: at low pH values the enzyme is inactivated slowly; there is presumably a metal involved in the inactivation, whose concentration certainly varies from laboratory to laboratory; and the previous reports dealt only with final values, not rates. In order to maintain maximal activity, a metal binder, a reducing agent, or both should be included at any pH.

The partially purified preparation acetylates a number of aromatic amines, as well as histamine, P-phenylethylamine, and glucosamine. No acetylation was observed when aromatic amines substituted with acidic groups in the ortho position were used. These compounds did not in- hibit the acetylation of other aromatic amines. The extent of purification is still too limited to indicate whether one or more enzymes are responsible for the different activities. Although our results with benzylpenicillin confirm those of Chou and Soodak in indicating differences in


http://ww

w.jbc.org/

Dow

nloaded from

http://www.jbc.org/


glucosamine and PNA acetylation, the mechanism of this inhibition is not sufficiently clear at present to require the conclusion that separate enzymes are involved. This is particularly true in view of the marked differences in the rate of acetylation of PNA and glucosamine.

The direct spectrophotometric method for following the acetylation of aromatic amine affords a convenient procedure for the measurement of acetyl CoA. Substrates can be selected to permit the reaction to be followed at convenient wave-lengths; the reaction may be followed spectrophotometrically by measuring either the disappearance of the free amine or the appearance of the acetyl compound. This method for the deter- mination of acetyl CoA has certain advantages over the other two spectrophotometric procedures available. Whereas the phosphotransacetylase reaction is followed at 232 to 240 rnp and the citrate reaction at 340 rnp, much higher wave-lengths can be used with aromatic amines. This permits reactions to be studied in crude systems and in the presence of other substances which absorb strongly in the ultraviolet. This method also permits acetylation to be measured in systems containing pyridine nucle- otides.

We wish to thank Mr. Frank Suggs for technical assistance during these studies.

SUMMARY

1. The acetylation of p-nitroaniline to p-nitroacetanilide affords a convenient spectrophotometric method for following acetylation reactions, as well as for the assay of acetyl CoA.

2. A partially purified pigeon liver preparation has been used to carry out this acetylation of p-nitroaniline, as well as the acetylation of p-aminobenzoic acid, histamine, and numerous other amines.

3. The acetylating enzyme has been shown to contain an essential sulfhydryl group.

BIBLIOGRAPHY

1. Lipmann, F., J. BioZ. Chem., 160, 173 (1945). 2. Kaplan, N. O., and Lipmann, F., J. Biol. Chem., 174, 37 (1948). 3. Nachmansohn, D., and Machado, A. I,., J. Neurophysiol., 6, 397 (1943). 4. Chou, T. C., and Soodak, M., J. Biol. Chem., 196, 105 (1952). 5. Millican, R. C., Rosenthal, S., and Tabor, H., J. Pharmacol. and Exp. Therap.,

97, 4 (1949). 6. Lynen, F., Reichert, E., and Rueff, L., Ann. Chem., 674, 1 (1951). 7. Mehler, A. H., and Knox, W. E., J. Biol. Chem., 187,431 (1950). 8. Gregory, J. D., Novelli, G. D., and Lipmann, F., J. Am. Chem. Sot., 74,854 (1952). 9. Wilson, I. Il., J. Am. Chem. Sot., 74, 3205 (1952).

10. Stadtman, E. R., and Kornberg, A., J. Biol. Chem., 203, 47 (1953).


http://ww

w.jbc.org/

Dow

nloaded from

http://www.jbc.org/


11. Tabor, H., and Mosettig, E., J. BioZ. Chem., 180,703 (1949). 12. Stadtman, E. R., and Lipmann, F., J. Biol. Chem., 186, 549 (1950). 13. Stadtman, E. R., J. Biol. Chem., 198, 527 (1952). 14. Ochoa, S., Stern, J. R., and Schneider, M. C., J. Biol. Chem., 193, 691 (1951). 15. Warburg, O., and Christian, W., Biochem. Z., 310, 384 (1941-42). 16. Moore, S., and Stein, W. H., J. Biol. Chem., 178, 367 (1948). 17. Willstatter, R., and Kraut, H., Ber. them. Ges., 68, 1117 (1923). 18. Chou, T. C., and Lipmann, F., J. Biol. Chem., 196, 89 (1952). 19. Stadtman, E. R., J. Biol. Chem., 196, 535 (1952). 20. Stern, J. R., Shapiro, B., Stadtman, E. R., and Ochoa, S., J. Biol. Chem., 193,

703 (1951). 21. Warburg, O., Ergeb. Enzymforsch., 7, 210 (1938). 22. Stadtman, E. R., Abstracts, American Chemical Society, 122nd meeting, At-

lantic City, Sept., 32C (1952). 23. Koshland, D. E., Jr., J. Am. Chem. Sot., 73, 4103 (1951). 24. Voegtlin, C., Johnson, J. M., and Rosenthal, S. M., Pub. Health Rep., U. S. P.

H. S., 46, 2234 (1931). 25. Hellerman, L., Chinard, F. P., and Deitz, V. R., J. Biol. Chem., 147, 443 (1943). 26. Rosenthal, S. M., and Tabor, H., J. Pharmacol. and Exp. Therap., 92,425 (1948). 27. Millican, R. C., Arch. Biochem. and Biophys., 42, 399 (1953). 28. Handschumacher, R. E., Mueller, G. C., and Strong, F. M., J. Biol. Chem., 189,

335 (1951). 29. Soodak, M., Abstracts, Division of Biological Chemistry, American Chemical

Society, 119th meeting, Boston, 18C, Apr. (1951). 30. Morgan, W. T. J., and Elson, L. A., Biochem. J., 28, 988 (1934). 31. Racker, E., and Krimsky, I., J. Biol. Chem., 198, 731 (1952). by guest on A

pril 3, 2018http://w

ww

.jbc.org/D

ownloaded from

http://www.jbc.org/

StadtmanHerbert Tabor, Alan H. Mehler and E. R.

AMINESTHE ENZYMATIC ACETYLATION OF

1953, 204:127-138.J. Biol. Chem.

http://www.jbc.org/content/204/1/127.citation

Access the most updated version of this article at

Alerts:

When a correction for this article is posted

When this article is cited

alerts to choose from all of JBC's e-mailClick here

tml#ref-list-1

http://www.jbc.org/content/204/1/127.citation.full.haccessed free atThis article cites 0 references, 0 of which can be


http://ww

w.jbc.org/

Dow

nloaded from

http://www.jbc.org/content/204/1/127.citationhttp://www.jbc.org/cgi/alerts?alertType=citedby&addAlert=cited_by&cited_by_criteria_resid=jbc;204/1/127&saveAlert=no&return-type=article&return_url=http://www.jbc.org/content/204/1/127.citationhttp://www.jbc.org/cgi/alerts?alertType=correction&addAlert=correction&correction_criteria_value=204/1/127&saveAlert=no&return-type=article&return_url=http://www.jbc.org/content/204/1/127.citationhttp://www.jbc.org/cgi/alerts/etochttp://www.jbc.org/content/204/1/127.citation.full.html#ref-list-1http://www.jbc.org/content/204/1/127.citation.full.html#ref-list-1http://www.jbc.org/

the enzymatic acetylation of amines the enzymatic

Documents