Oswald Avery identifies nucleic acids as the active principle in bacterial transformation
Date: 2015-10-07; view: 478.
The molecular weight of nucleic acid was found to be much higher than the tetranucleotide hypothesis required, but it was still viewed as a uniform polymer, like starch, unaffected by its biological source
In 1940, Avery once again focused on the problem of bacterial transformation. After Colin MacLeod left to join the faculty at New York University later that year, Maclyn McCarty joined the laboratory in 1941 and aided Avery in this research. Avery and McCarty focused first on purifying the transforming substance. Using refined versions of Colin M. MacLeod's preparation techniques, Avery and McCarty isolated biologically active "transforming principle" from samples of pneumococci. After jump-starting the research, Avery was increasingly preoccupied with the step-by-step purification of the transforming agent and its identification. Initially, transformation had been a tentative and delicate phenomenon that was difficult to consistently recreate. Avery later told Rollin Hotchkiss, "Many are the times we were ready to throw the whole thing out the window!" Eventually, Avery and McCarty were able to take a culture of pneumococci of an R form that had been attenuated from an S of Type II over the course of thirty generations, and add to it to the highly purified deoxyribonucleic acid (DNA) extracted from an S of Type III. This process resulted, by the next generation, in large and fully developed colonies of S Type III, which remained stable through several generations. After achieving reliable and long-lasting transformation, Avery turned to prove that it was caused by DNA alone, despite the prevailing conviction of most geneticists, and even his own earlier belief, that DNA was a simple molecule and that genes must be composed of protein, a seemingly more complex substance
As Avery and McCarty turned their attention to the chemical analysis of transformation, they found that proteases (enzymes that deactivate proteins) and lipases (enzymes that destroy lipids) did not inactivate the transforming principle, and thus concluded that the substance was essentially protein- and lipid-free. However, it was not a carbohydrate like the polysaccharide capsular material, as carbohydrates are not precipitated by alcohol, as was the "transforming principle." They soon determined that the substance was rich in nucleic acids, but ribonuclease, an enzyme that destroys ribonucleic acid (RNA), did not inactivate the substance either. Further, the transforming substance had a high molecular weight, as did DNA, and gave a strong reaction to the Dische diphenylamine test, which detects for the presence of DNA. Avery and McCarty concluded that the transforming substance, which produced permanent, heritable change in an organism, was DNA.
(1944)
"If the results of the present study of the transforming principle are confirmed, then nucleic acids must be regarded as possessing biological specificity the chemical basis of which is as yet undetermined."
A few microbiologists and geneticists had taken an interest in the physical and chemical nature of genes before 1944, but the Avery–MacLeod–McCarty experiment brought renewed and wider interest in the subject. While the original publication did not mention genetics specifically, Avery as well as many of the geneticists who read the paper were aware of the genetic implications—that Avery may have isolated the gene itself as pure DNA. Biochemist Erwin Chargaff, geneticist H. J. Muller and others praised the result as establishing the biological specificity of DNA and as having important implications for genetics if DNA played a similar role in higher organisms. In 1945, the Royal Society awarded Avery the Copley Medal, in part for his work on bacterial transformation.
(1950)