Little is known about the abundance or function of small proteins in any organism. These proteins, defined as those containing fifty or fewer amino acids, are difficult to isolate and identify using standard biochemical techniques, and thus are commonly missed in protein identification studies. In addition, it is difficult to identify the short genes that encode small proteins using bioinformatics. As a result, the small proteins are significantly underestimated. The PI's research and that of others has shown that bacteria synthesize many more small proteins than previously thought, and the total size of the small protein proteome in E. coli or any other organism is still unknown; but is estimated to be in the hundreds. To identify new small proteins in E. coli, the investigator plans to biochemically assay for protein synthesis from candidate small open reading frames in cells grown under different growth and/or stress conditions. This research will help to elucidate the true number of small proteins in the E. coli proteome, as well as provide an initial characterization of protein regulation and biochemical interactions with other proteins that will serve as a foundation for identifying the function of these proteins in the future.
There are many aspects of the small protein identification project that make it ideal to expose undergraduate students to a hypothesis-driven, inquiry-based molecular biology learning experience. Teams of students will use basic bioinformatics to identify small open reading frames that potentially encode small proteins. These students will test their hypotheses in a follow-up molecular biology laboratory course. Ultimately, information gained from this project on small protein abundance, growth- and stressÂ-dependent accumulation and physical interactions will provide a critical basis for offering interested Towson students additional opportunities for conducting independent research in the PI's laboratory.