PC1 β-lactamase

Protein Data Bank The PDB is a resource which compiles essential information about many proteins. Go to the PDB website at www.pdb.org.

PC1 – β-lactamase We’ll be investigating β-lactamase. Type PC1 into the search box. More than one result is produced… How should we choose the one we want to look at? What does the information given about each mean? How do you know? Which one do you think we should choose?

3blm Click on 3blm Underneath the image of the structure, click on ProteinWorkshop. Ribbon vs. Atom view Color changes Label changes

Our preferred view Ribbons Colored by compound No water molecules visible (for now)

1°, 2° Structure Divide into pairs. Each group will investigate the following portions of the enzyme. Residues 31-100 Residues 101-170 Residues 171-240 Residues 241-290 First make your residues visible as atoms and molecules (visibility tool) As you go, use your amino acid reference translate each residue’s 3 letter abbreviation into it’s 1 letter abbreviation. Type this your 1 letter list into a Powerpoint slide and email it to me. We’ll combine this information in the next slide. Use the labeling tool to label your segment by residue. Prepare to show and tell and to describe the geometry of your segment

1° structure Residues 31-100 KELNDLEKKYNAHIGVYALDTKSGKEVKFNSDKRFAYASTSKAINSAILLEQVPYNKLNKKVHINKDDIV Residues 101-170 AYSPILEKYVGKDITLKALIEASMTYSDNTANNKIIKEIGGIKKVKQRLKELGDKVTNPVRYEIELNYYS Residues 171-240 PKSKKDTSTPAAFGKTLNKLIANGKLSKENKKFLLDLMLNNKSGDTLIKDGVPKDYKVADKSGQAITYAS Residues 241-290 RNDVAFVYPKGQSEPIVLVIFTNKDNKSDKPNDKLISETAKSVMKEF

2° structure

Hydropathicity Describes the hydrophillicness / hydrophobicness of a protein. Two ways: Change coloring in Protein Workshop Input your sequence into a Kyte-Doolittle converter (http://www.vivo.colostate.edu/molkit/hydropathy/index.html) Do both of these and be prepared to describe your segment’s 2° structure in terms of the result.

Hydropathicity

Hydropathicity

3° structure Let’s try to group the types of tertiary indicators found in your segments α-helices 32-40, 69-81, 119-129 , 132-142, 145-154, 183-194, 201-213, 221-224, 277-287 β-sheets (43-50, 56-60, 94-96) (230-237, 244-251, 259-256) Turns 51-53, 62-63, 91-92, 107-108, 113-114, 155-156, 166-167, 174-175, 178-179, 194-195, 218-220, 227-228, 241-242, 253-254, 270-271, 288-289 Folds – α-helix between domain 1 (β-sheet 5 antiparallel strands and 3 α-helices) and domain 2 (α-helices.)

3° structure

4° structure With your partner, go back to the pdb mainpage for 3blm and try to determine the 4° structure. Look to see if there are any ligands. Think about what a ligand would be and would mean in this context. Prepare to justify your conclusions to your classmates.

4° structure No 4o structure. No ligands.

Remember what β-lactams are supposed to do…

What about water in the β-lactams scenario? It is this reaction which β-lactamases catalyze!

So what now? You’re the drug researcher trying to develop a effective β-lactam… What considerations should you investigate regarding β-lactamases? What traits of the β-lactams? What traits of the β-lactamases? Which are more easily studied?

References Buckwell, S. C.; Page, M. I.; Longridge, J. L., Hydrolysis of 6-alkyl penicillins catalyzed by b-lactamase I from Bacillus cereus and by hydroxide ion. Journal of the Chemical Society, Perkin Transactions 2: Physical Organic Chemistry (1972-1999) 1988, (10), 1809-13. Herzberg, O., Refined crystal structure of beta- lactamase from Staphylococcus aureus PC1 at 2.0 A resolution. J Mol Biol 1991, 217, (4), 701-19. 3. www.pdb.org 4. Protein Workshop Viewer accessed via www.pdb.org