Chem 8157

Lecture 1,2 (2/28)
1) We are unsure of the mechanism of iodoacetate (Manz Fig. 7.8). We have reason to believe it could be SN2 due to the good iodide leaving group but we would like clarification.

ANSWER: I recall it is an SN2 reaction.

2) What is the purpose of glycine in 2D SDS-PAGE? We belive it is an additive.

ANSWER: At the working pH it is a zwitterion and buffer. It helps maintain the pH, and conductivity.

3) CNBr reacts with methionine but what prevents it from reacting with alkylated cysteine residues?

ANSWER: I do not know. People usually do not alkylate cysteine residues if they will use CNBr.

Lecture 2,3 (3/10, 3/11)
1) Feel free to not respond to this question: should we memorize the alternative fragment ion definitions? (yo, y*, y++, etc.)
ANSWER: Learn those that were included in exercises, presentations, and lecture notes.

2) When sequencing with MS/MS, why don't you see the terminal y/b ions? Reference 11. We understand how to determine the missing ions but why aren't they observed in the first place.
ANSWER: Mass analyzers have different transmission at different m/z ranges. This transmission is very poor at the low m/z range. Thus, the y1, y2, b1, b2, etc. ions may be produced by CID but they are not transmitted by the mass analyzer.

3) What is the difference between monoisotopic mass and average mass? Is monoisotopic mass the mass of the specific ion being used? Is average mass the average isotopic mass of the element found in nature?
ANSWER. Monoisotopic mass: for each atom in the formula use the mass of only one isotope. Average mass: for each atom in the formula use the average mass determined by the isotopic abundance of the element.

4) In slides 21-22, parts c-f, what is the purpose of the first, unlabeled quadrupole (the quad that is shown to the left of Q1)?
ANSWER: Although it is represented as a quadrupole, it may represent an electromagnetic lens. This lens may help focus ions, homogenize their kinetic energy, or to eliminate ions that do not belong to the ions of interest.

5) In the FT-MS and LTQ-FT, what's the purpose of the magnets?
ANSWER: Both instruments rely on ion cyclotron resonance. The magnetic field produced by the magnet causes an electrical force on a moving charged ion. This electrical force is counterbalance by a centripetal force produced by the orbiting ion (perpedicular to the direction of the magnet). This defines a stable trajectory at a given frequency for a given m/z value. This frequency of rotation is captured by a sensor (induced current) that is used then to identify the m/z value.

6) For slide 28, parts B and C, we understand that these are diagramms for determination of of phosphorylated and glycosylated proteins. What is the reason for the MS/MS then if all that is being examined is the presence of modified proteins (not peptides)?
ANSWER: Peptides, not proteins are being examined. One detects the glycosylation (part b), or loss of phosphoric acid mass (part c) in peptides with these modifications.

Lecture 5/6 (3/24)
1) In the exponentially modified PAI equation (emPAI = 10^PAI -1), why is the -1 term added?
ANSWER: Good question. My guess is that when there are no peptides PA1 = 0; when this happens, emPAI must be zero.