Week 3: June 3 2016

Friday, 6/3
Members present: Hamed, Kat, Zarifah, Alex, Bohdan, Tam
Lab Stuff: (research, plasmid design, protocol design, troubleshooting)
Lab moving day
GolS cleaved His (Ser): S277165
Results suggested GolS cleaved His (Ser) indicated a greater deviation from wt GolS and GolS cleaved His (Gly).
wt GolS.gif
GolS cleaved His (Ser).gif
The alpha helix near the C-terminus elongates and appears to be drastically different from wt GolS. There is a suspect that the Ser155 residue forming H-bond with other residues at the N-terminal DNA binding domain.
This is a big indication that any polar residues can change the conformation of the helix-turn-helix structure (Ser, noncleav His-tags). His-tag is not recommended to be done with the Serine version at the end.
Despite the cleavage of the linker, there is a small change in the pocket binding site for metal ions and we are skeptical on whether it would greatly affect the metal ion binding sites.
Images of the the wt GolS and GolS + Gly155:
wt GolS pocket size.PNG
The pocket size on the wt GolS is very tiny and almost hard to visualize on the program.
GolS + Gly155 pocket site.PNG
The pocket size on GolS with extra Gly155 residue is larger and suspected to be less specific to Au ions. The pocket may show certain affinity to other ions besides Au ions.
In conclusion, it is recommended that wt GolS should not be used with a linker His-tag as the binding site is heavily sensitive to even a slight change in the residue addition or point mutation.
Looking for other alternatives to purification of GolS.
Looking into Rosetta with the dry lab.
Administrative: (shipment orders, inventory)
Email Updates: (Correspondence with professors, shipping companies, etc)
Emailed Ms Gorette Silva about WB407 room bookings for June 6th (with Mr Caffrey at 3PM) and June 8th (for Dr Papangelakis at 12 noon) for 2 hours - room available but pending confirmation
Contact to the provider of heparin-sepharose column (GE Health Life Sciences) and confirm on the date in which the column will arrive.
Meeting Notes: