University Honors

Escherichia coli cells can use up to 40% of their energy for making ribosomes (Bubunenko et al., 2006). Ribosomes are important for converting the genetic information encoded in mRNA to protein. As a result, ribosomes can directly play a critical role in the cell’s response to environmental changes via proteins. The ribosome is a complicated machine made up of many components of ribosomal RNA (rRNA) and additional proteins that join to form an active, mature 70S ribosome. The ultimate formation of mature 70S requires the help of factors, known maturation factors, such as RimM. Previous research has shown that Cryogenic Electron Microscopy (CryoEM) structures of ribosomes isolated from E. coli cells missing RimM are distinct from the wildtype that contains RimM (G. Demo, Personal Communication)(See Appendix A Figure 1). The removal of maturation factors such as RimM may have implications in forming the mature 70S ribosome. If the mature 70S ribosome is not formed or does not function properly, this can affect the translation mechanisms in the mutant. G. Demo, our collaborator, believes that the ΔRimM mutants contain a prominent additional unknown density that has not been identified before from the wildtype cells. This additional density is thought to be a protein with a known molecular weight of 40-50 kDa and has been hypothesized to be located on the 70S ribosomal subunit and seems to assist in the maturation of Helix 68 (H68) of the ribosomal RNA (G. Demo, Personal Communication). H68 is an important element during protein synthesis that is involved in ribosomal movement during protein elongation by switching between two conformations, so alterations in the H68 function may also have implications with the ribosome machinery (Cimicata et al, 2022). I investigated whether or not the unknown protein exists in the 70S ribosomal subunit in the presence (WT) and absence (i.e., knock-out, KO) of RimM from both MG1655 and MRE600 E. coli strains by isolating and purifying the 70S subunits of the ribosome using sucrose gradients.