BASIL Biochemistry Curriculum 2019

Your students will use a computer alignment algorithm that includes quantitative measures (Levenshtein distance and RMSD values) and visual inspection to measure quality of alignments.

These results can be compared and contrasted with sequence alignment (BLAST, Pfam) and and full backbone alignment (Dali) results in other BASIL modules.

This module requires that you to install PyMOL and ProMOL on student-accessible computers and requires internet access.

Protein BLAST searches identify sequence similarity between a query protein and a database of other proteins.

Sequence similarity does not guarantee similarity in function.

Highly conserved amino acid residues does not guarantee similarity in function.

Dali searches identify backbone structure similarity between a query protein and 3D structures of proteins from the PDB.

Backbone structure similarity may not guarantee similarity in function since side chains are important for function.

Noncovalent interactions stabilize the binding of ligands to macromolecules like proteins.

Ligand binding is complex; chemical modifications to a ligand may not result in changes to the interaction energy due to competing stabilizing and destabilizing interactions.

Introduction of a foreign gene into E. coli allows researchers to express proteins of interest.

Protein expression can be controlled using knowledge of operators and repressors in E. coli.

Purification of proteins is usually accomplished by lysis, removal of nucleic acids, and affinity chromatography utilizing a 6-histidine tag or fusion protein tag.

Measuring protein concentration is important for determining protein purification yield and amounts to use in further analyses.

To carry out this lab students need vertical electrophoresis equipment.

The students can use pre-purchased gels or pour their own.

Activity assays with a broad substrate can be used to identify the class of enzyme while specific substrates can yield more information about enzyme function.

Determining kinetic parameters for the enzyme of interest can indicate the best substrate(s) for the enzyme and consequently provide insight into its function within an organism.