Molecular cloning of recombinant DNA : proceedings of the Miami winter symposia, January 1977 / edited by W.A. Scott, R. Werner ; sponsored by the Department of Biochemistry, University of Miami, School of Medicine, Miami, Florida.

Date:
1977
Catalogue details

Licence: Attribution-NonCommercial 4.0 International (CC BY-NC 4.0)

Credit: Molecular cloning of recombinant DNA : proceedings of the Miami winter symposia, January 1977 / edited by W.A. Scott, R. Werner ; sponsored by the Department of Biochemistry, University of Miami, School of Medicine, Miami, Florida. Source: Wellcome Collection.

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    6 PAUL BERG I learned two important lessons from that experience: To be concerned with the purity of enzjmies when examining their reaction mechanisms; and, that irrespective of whether the hypothesis or model being tested is correct, there is a strong likelihood of making an original finding if the experi¬ ment being done has never been done before. A Return to Reality Although our departure from Copenhagen was a sad one we were consoled by the fact that it had been a happy and success¬ ful year; besides, there were new things to learn and new worlds to conquer. During my year abroad Romberg had moved from the NIH to establish a new Department of Microbiology at Washington University Medical School. When I arrived during November 1953, he eagerly showed me where I would work - one of the few refurbished laboratories in the Department - and even before unpacking my bags, we were discussing what problem I would tackle. He suggested that I could pursue his earlier observation of the conversion of phosphoryl choline to phospholipid (9) or take on some aspect of the work he and Irving Lieberman were doing on pyrimidine nucleotide formation. But I had been intrigued by a report earlier that year (10), from Lipmann's and Lynen's laboratories, that apparently solved the mechanism of acetylCoA formation by aceto-CoA kinase. Although it was known that the condensation of acetate and CoA was accompanied by the cleavage of ATP to AMP and PPi, the mechanism of this ternary reaction was a mystery. The joint Boston-Munich communique proposed three partial reactions to account for the overall process (Figure 2), because their enzjnne prepara¬ tion alone catalyzed the exchange of PPi with the pyrophos- phoryl group of ATP, as well as the exchange of CoA-bound acetyl groups with free acetate. What attracted me about their model was the proposal of an AMP-enzjrme intermediate and particularly the possibility that analogous nucleotidyl-enzyme compounds, involving other ribo- and deoxynucleotides, might be substrates in polynucleotide synthesis. But Romberg was dubious about this possibility; moreover, he felt that the Lipmann-Lynen results could be attributed to contaminating enzymes that catalyze a reversible pyrophospho- rolysis of NAD and FAD, thereby, incorporating isotope from PPi into ATP. Nevertheless, because it seemed important to establish or demolish this model of acetylCoA synthesis, I persuaded him to let me try to isolate the putative adenyl- enzyme compound.