In Vivo Pathways of Molecular Evolution: Acquisition of Thermostability by Mesophilic Adenylate Kinases

William Marsh Rice University, Houston TX

Investigators

Abstract

The principles of evolution are well known and are reflected in the beauty and complexity of life on this planet. Understanding how life evolves and changes through time is an important question in modern biology. Even before DNA was discovered to be the hereditary material of life, the ideas underlying evolution could be observed readily by the domestication of plants and animals. Perhaps one of the most well known examples of domestication is the dog, once a wild wolf and now a companion. Though it is easy to appreciate the outward differences between a dog and a wolf, it is more difficult to determine how small changes in the genes and the proteins encoded by them give rise to changes in the characteristics of each breed. The PI studies the evolution of molecules and how changes in environmental conditions such as temperature select for changes (mutations) that make those molecules better adapted. Typically, the adaptation of organisms to new conditions takes much longer than is practicable in lab studies. By focusing on a simple thermophilic organism (a bacterium that lives at high temperatures) that reproduces in 30 minutes, thousands of generations can be studied during a single laboratory experiment. To facilitate these studies the ''weak link'' approach was developed with previous NSF support. In the ''weak link'' approach, survival of a genetically modified thermophilic organism (Geobacillus stearothermophilus) is dependent on changes to an essential molecule that only functions at lower temperature. In order to reproduce at higher temperature, mutations must occur to the essential gene. By growing the bacteria at low temperature and then gradually increasing the temperature the bacteria can be quickly adapted into one that can live at high temperatures. It is then possible to determine how changes to the essential gene produced a protein that was well adapted to work at high temperature and permit the bacteria to survive in the new environment. Experimental evolution of a single gene also provides an opportunity to investigate the functional intermediates of protein adaptive evolution during the course of natural selection. Several fundamental questions can be addressed through this work. For example: How reproducible is evolution? If the experiment is performed three times do the same changes occur and in the same order? What happens if the conditions of the selection change? How will adaptation change the molecule in response? In addition to exploring the basic principles for all life, these findings have application to protein engineering and biotechnology for the production of more rugged proteins. Broader Impact The integration of science education and research is a primary focus of the investigator''s laboratory. Nine undergraduate and three graduate researchers have carried out work on this project as independent research and have presented findings at scientific conferences and in published research. The breadth of the project from organism to electron density in high resolution structures requires students to become broadly educated in evolution, microbiology, biochemistry and biophysics. The PI is faculty mentor for the NSF-IGERT Program for Cellular Engineering, Houston Molecular Biophysics Program and the Gulf Coast Consortia and is a co-founder of the Rice Center for Evolution. The PI teaches Biochemistry and incorporates the lessons learned from these experiments into the discussion of protein folding and stability as well as how evolution impacts our world through the rise of drug resistant pathogens. In the last three years, the lab has hosted 7 High School groups (including a High School AP teachers program) as well as mentoring summer researchers from the under represented Hispanic community through the Rice AGEP program. The PI has also used this project as the basis for community based seminars on the nature of evolution (''Evolution in a coffee pot'').