Journal Club 10 take home questions

Here is a link to the take home questions to be completed before next Journal Club:

https://docs.google.com/viewer?a=v&pid=explorer&chrome=true&srcid=0B_Gl3lMyhDsoZjM0ZWUxMGUtMGFkMi00N2VhLWI3MmEtMjQ3NWI2ZDdmOThj&hl=en&authkey=CPHfi9QH

 

Quiz questions – Instructions: Answer questions on sheet individually and turn in during class

Q1:  What disease are they trying to treat? Why is it particularly hard to target?

Answer:

Q2: What is the purpose of this K* program? To get a better feel for the big picture, it is sometimes helpful to read the Conclusions section first.

Answer:

Q3: What does WT stand for in the paper? e.g. in Table 1:

Answer:

Q4: What does the Negative vs Positive selection mean?

Answer:

Q5: In the Determination of Crystal Structure section, what type of interaction is lost when Phe92 is changed to Ile? What property or feature of F vs I would explain this loss?

Draw a diagram of F and of I.

Then circle the F92 in Fig.1

Answer:

Q6: When an enzyme in bacteria mutates and loses specificity for the drug, why doesn’t it lose its specificity for the substrate too? Aren’t most drugs supposed to be really similar to the substrate?

Answer:

Q7: In table 3, which mutant has the highest affinity for the substrate? What piece of evidence helps to support this?

Alternatively, in Table 4, which mutant is least affected by the inhibitor (Compound 1)?

Answer:

Q8: In the M&M section –they say that out of the 10 active site residues they would mutate 2 while letting the other 8 sites change rotamers. Why not allow all 10 to be mutated?

Answer:

Q9: What was the chromophore (thing that absorbs light) in their enzyme assay? What wavelength (is this in the visible or not)?

Answer:

Q10: They chose MRSA – if you could choose another organism to apply this research to, what would you choose?

Answer:

Q11: They don’t really do any virtual screening here. How could we apply this research to our stream?

Answer:

Journal Club #10

Here is the final journal club article.

You need to download both the Main article and the Supplementary info (PDF with more graphs and tables)

Go to PubMed to get it.

http://www.lib.utexas.edu/indexes/titles.php?id=299&p=print

If you are off campus, use the VPN to connect to the campus network.

http://www.utexas.edu/its/vpn/index.php

Frey, K. M.; Georgiev, I.; Donald, B. R.; Anderson, A. C., Predicting resistance mutations using protein design algorithms. Proc Natl Acad Sci U S A 2010, 107 (31), 13707-12.

Next Meeting – changed to Monday the 22nd

POTENTIAL CHANGE

Since we cannot get the room at the original time, we would like to change the meeting to 4:00 – 5:00 on Monday.

If you CANNOT make that time – email Dr. B

We are hoping to have it in WEL 4.238 conference room.

Thanks,
Dr. B

Journal Club #9

Here is the next paper for Journal Club that Gabriela will present.

Evers, A.; Klabunde, T., Structure-based drug discovery using GPCR homology modeling: successful virtual screening for antagonists of the alpha1A adrenergic receptor. J Med Chem 2005, 48 (4), 1088-97.

And the Quiz questiosns

Journal Club #8 Quiz Answers

Q1: If all of the proteins are made on one long strand, how will they be able to function within the cell?

Answer: they are cleaved during post-translational modification by the protease

Q2: Why aren’t the structural proteins good targets of Dengue infection?

Answer: They are good – maybe not as critical for replication of the virus but needed for viral fusion and entry

Q3: Where does the protease cleave the viral protein?

Answer: where the arrows are in Figure 5

 

 

Q4: For the DDD-T, why do they do two phases? What purpose does each phase serve?

Answer: virtual screening in first phase gets poses, then a top amount of these are passed on to phase two for a more rigorous SM-FEB free energy of binding calculation using Structural Mechanics. This gives a more accurate energy score which can then be used to ‘cherry pick’ the best candidates

Q5: What are the advantages and disadvantages of distributed computing to cluster computing?

Answer: shared bandwidth is low, many different operating systems and codes, cheap, easy to maintain

Journal Club #9

Quiz Questions for Journal Club:

https://docs.google.com/viewer?a=v&pid=explorer&chrome=true&srcid=0B_Gl3lMyhDsoODU5ODlhNzctNjdjYS00MzZmLTgwOTQtMTBlMTJlMDc3Mjgx&hl=en&authkey=CIadvdcN

Here is the paper for next Tuesday on malaria

https://docs.google.com/viewer?a=v&pid=explorer&chrome=true&srcid=0B_Gl3lMyhDsoYjg0OTQ0OTgtOWQ2Yy00MzllLWEyYWQtM2I4NTNiNDNiMjM3&hl=en&authkey=CJG78X4

And supporting Info

https://docs.google.com/viewer?a=v&pid=explorer&chrome=true&srcid=0B_Gl3lMyhDsoMzUyMTlmZTUtYzJjZS00ZTc5LWJiZWMtYzNlM2RhNmNlZDAx&hl=en&authkey=CLLo1OYN

Journal Club #8

Here is the paper for Tuesday, with questions to follow later:

https://docs.google.com/fileview?id=0B_Gl3lMyhDsoYzA2ODBmYTktMTRlMi00NjMyLTlmN2QtNzM1MzI4Y2EwMDNi&hl=en&authkey=CNHK4KcN

Journal Club 7 Study Question Answers

It says Ricin is not infectious – but this is used by bioterrorists – don’t they use infectious agents?

Answer: not a replicating organism – more of a poison

What is the mode of action for ricin and what does it carry out this action upon?

Answer: depurinates (removes a purine – adenine) form 28s ribosomal RNA

What is STxA1 and how is it different than Ricin?

Answer: shiga toxin. They express both in this paper

In the intro – what do they say is the advantage of the PBA ligand?

Answer: more soluble and more advantages for inhibitor design

How many compounds passed the filter in their library?

Answer: 306  – same as cb306 library we use for screening

Why did they not use a His tag?

Answer: poor expression levels

What is a French press – a Francophile’s offensive strategy in basketball?

Answer: machine for cell lysis

What is a typical yield for their protein? Do you get the same amount?

Answer: 5mg per liter

How does their inhibition assay work? Is this different from an enzyme assay we would do in our research?

Answer: monitor luciferase bioluminescence after the inhibitor slows RTA’s ability to stop the ribosome

What do they dissolve their compounds in for the assay? To what final percentage?

Answer: DMSO to 1%

What type of signal is measured for the cell assay? Could we do this in our lab?

Answer: luminescence as well

What does Fig 2 tell us?

Answer: that scores don’t correlate

In section 3.3 – it says that the IC50 curve reflects the limited solubility of the compound. What should a normal IC50 curve look like?

Answer: sigmoidal – add more should shut it down

In Figure 4, which compound is more cytotoxic alone?

Answer: left one

In Table 1, how can the compounds inhibit RTA but not inhibit RICIN?

Answer: enzyme assay vs. cytotoxicity assay – can’t get into cytoplasm

Journal Club 6 quiz answers

What is a promiscuous inhibitor? Does that mean it should be tested for STD’s?

ANSWER: non specific inhibitor – tends to inhibit many enzymes. But usually does not have high affinity and will not lead to a drug

While the inhibitors can be promiscuous, it says the CAII enzyme is just a boring old housekeeper. What does this imply?

ANSWER:it is responsible for carrying out normal functions of homeostasis and is important – don’t want to inhibit unless necessary.

What is glycerol in this paper?

ANSWER: cryoprotectant

What is so special about the binding of the inhibitors which they are studying? Haven’t these been documented in the literature before?

ANSWER: yes have been studied as individuals but not as a linked sulfonamide+coumarinyl structure. So, it turns out to have two unique binding mechanisms – to the Zinc and also one just occludes the whole active site.

Based upon Table 1., which one would be good for use as a treatment for anti-malarials?

ANSWER:none – AZA has good Ki but would have bad side effects on the human isoforms because Ki are too good there and it is bad to inhibit CA2 because of its housekeeping role in human cell.

What do the blue and orange represent in Fig 3? Is this odd to you?

ANSWER: hydrophilic vs hydrophobic side of active site. Usually not oriented longitudinally like this but more often have ‘pockets’ of hydrophobic or hydrophilic

Near the end of the paper, why specifically do they say Inhibitor #1 is good at inhibiting hCAI but not hCAIX and also hCAXII?

ANSWER: PHE 131 is substituted by other amino acid – so the coumarin ring cannot stack against the subsitutions

In what way is Phenol red used?

ANSWER: as an indicator of pH, spectrophotometrically measured

They mention that their protein was ‘induced’ – does this mean it is going into labor?  What is going on in those steps and what is IPTG and what is its purpose?

ANSWER: IPTG binds to promoter region of GOI and drives transcription of the gene which leads to translation of the protein

Journal Club 7

Here is the paper:

https://docs.google.com/fileview?id=0B_Gl3lMyhDsoYzVkOTVjZjYtOGZmZS00ZGNmLWJkNDYtZjkyOGQ0NzRiZWVm&hl=en&authkey=CKWc488D

Here is the text below as a Word doc:

https://docs.google.com/fileview?id=0B_Gl3lMyhDsoMzUxOGM1ZmYtNmQzNS00YmU4LWI4YTMtYTljYzc2MWUyNDg5&hl=en&authkey=CPmJ5YsL

Paper 1 – presenter   Thao Pham

Bai, Y.; Watt, B.; Wahome, P.; Mantis, N.; Robertus, J., Identification of new classes of ricin toxin inhibitors by virtual screening. Toxicon 2010, 56 (4), 526-34.

Print out and turn in your paper. The quiz will consist of grading your notes and annotations on your printout of the journal paper. Essentially, read the paper and make notes and comments to yourself. Underline sentences that are important and make a few sketches or diagrams when useful.     You will be graded on the thoroughness of your note- taking.

Also, there is one PyMol exercise that will be graded: download the 1BR6 closed form of RTA and the 1IL5 open form. Align them to each other in PyMol. Show as ribbon. Find the two different Tyrosine 80’s (one for each PDB) and show as sticks. Make an image of how different the Closed vs. Open form is and turn it in with your Journal Paper.

Below are some study questions to think about for the paper:

It says Ricin is not infectious – but this is used by bioterrorists – don’t they use infectious agents?

What is the mode of action for ricin and what does it carry out this action upon?

What is STxA1 and how is it different than Ricin?

In the intro – what do they say is the advantage of the PBA ligand?

How many compounds passed the filter in their library?

Why did they not use a His tag?

What is a French press – a Francophile’s offensive strategy in basketball?

What is a typical yield for their protein? Do you get the same amount?

How does their inhibition assay work? Is this different from an enzyme assay we would do in our research?

What do they dissolve their compounds in for the assay? To what final percentage?

What type of signal is measured for the cell assay? Could we do this in our lab?

What does Fig 2 tell us?

In section 3.3 – it says that the IC50 curve reflects the limited solubility of the compound. What should a normal IC50 curve look like?

In Figure 4, which compound is more cytotoxic alone?

In Table 1, how can the compounds inhibit RTA but not inhibit RICIN?

Journal Club 6

VDSers,

Here is the paper and Take home quiz questions for this week’s journal club by Mike.

https://docs.google.com/fileview?id=0B_Gl3lMyhDsoNDk5NTlkZDItYjM1YS00NzcyLWJhMGEtNjEwNTk3ZmNiOTgw&hl=en&authkey=CL3SjccE

 

https://docs.google.com/fileview?id=0B_Gl3lMyhDsoNTg2NGJiOTAtZGU1My00MWZlLWJlZGEtYWU3MDQyNjZmMTYz&hl=en&authkey=CJ2HiskI

Journal Club 5

For the next journal club, there will be a Take Home Quiz instead of an in-class Quiz.

Here is a link to the Quiz

And  link to the Paper

Undergraduate Research Journal

A good place to publish your VDS work.

Undergraduate Research Journal information session announced
Description: Join the staff of the Undergraduate Research Journal for an
informal information session. The discussion will cover submission deadlines,
types of articles accepted, what we look for in choosing an article for
publication, the time line of the selection process and awards given for the
best articles.
tomorrow (Tuesday, October 12)

Time: 5-6:30 p.m.
Location: University Teaching Center 3.124
Admission: Free
URL: http://www.utexas.edu/research/student/urj/

_______________________________________________________

The Undergraduate Research Journal is a student-edited, multi-disciplinary
journal of undergraduate research at the University of Texas at Austin. Copies
of previous editions are available in the College of Natural Sciences’ advising
offices. Please send any questions regarding the URJ to texasurj@gmail.com.

Making a spec

So, now when the Red Tide specs break down – maybe we can just tap into our cell phones!

http://www.wired.com/gadgetlab/2010/10/in-high-school-chem-labs-every-camera-phone-can-be-a-spectrometer/

Journal Club 4

The new journal article has been uploaded to GDocs. Print it, read it 2x, and bring it to class.

https://docs.google.com/fileview?id=0B_Gl3lMyhDsoYTU1Nzc3ZGUtZGQwYS00Njg5LTg2ZjYtMjAxNjRjZTBjMzVm&hl=en&authkey=CI-5gNEB

PS: if you want to ask others in the group about the paper before class – this is a good place to post questions and answers in the comments after these posts.

That may better prepare you for the quiz questions.

Study Questions:

1. What EC number for this enzyme?
2. What is peptidoglycan and why is it important to bacteria? Do you have it in your body?
3. What is special about the ‘D’ in D-Ala-D-Ala ?
4. What two methods were used to assay efficacy of these inhibitors? What are the advantages of each over the other?
5. What is missing in Figures 3b,c, and d ?
6. Why do they test both E. coli and S. aureus?
7. What would be a next step after this research?