Bioinformatic Characterization of 3L1W

by Diana Severineanu (VI)

3L1W is one such protein selected from the Protein Data Bank (PDB) with an unknown function, but a known structure predicted to be a hydrolase. Using the BLAST and Pfam programs, which compare proteins based on amino acid sequences, the top results were all shown to be part of the endonuclease/exonuclease/phosphatase enzyme class, giving 3L1W a predicted EC number of 3.60.10.10. This EC number would later be used in the program ProMol to compare possible active sites. The DALI program, which compares proteins independent of sequence and based on 3D structure, was also used and returned similar results to BLAST and Pfam. In vitro, BL21(DE3) Competent E. coli cells were successfully transformed with the 3L1W plasmid, and the protein was expressed using OvernightExpression autoinduction medium. Cells were lysed and protein was purified using buffers of our design, and results were visualized using SDS-PAGE gels. Due to the disruption of the 2020-2021 school year due to the COVID-19 pandemic, I could not fully confirm our proteins’ predicted functions. This year, I aim to continue to determine the exact function of 3L1W by using various assays to test substrate affinity, rate of activity, and more, as well as model the reaction computationally and gain a deeper understanding of protein structure and its strong correlation with function.

Figure 1: A model of protein 3L,W from the Protein Data Bank