Take-Home Questions

Question 1

Briefly describe the major mechanisms that generate the vast majority of Abs; which property of the Abs that leads to its selection

Answer 1

Somatic hypermutation–> per variable region per cell division, there is one mutation in the antibody–>very high rate of mutation, leads to a slightly altered Ag-binding region
V(D)J recombination–> there are various V and J genes (and D on the heavy chain)–> differently joining can produce different variable regions–>e.g. have 10 Vs, 4 Js–> 40 different ways to combine–> 40 different Abs
–>also, there are different Ab genes to begin with
–>affinity leads to Ab selection

Question 2

Name and describe the 3 types of phagy, in 3 sentences or less

Answer 2

Macrophagy: machinery initiates the formation of a phagophore; phagophore engulfs the molecules–>becomes autophagosome; teh autophagosome then fuses with a lysosome to become an autolysosome, where degradation of the molecules will take plac
Microphagy: lysosome directly engulfs and degrades the substances, without the need of a phagophore
Chaperone-mediated autophagy: chaperone binds the protein, translocates it to the lysosome, where it is translocated through the lysosomal membrane and degraded

Question 3

Describe Dr. Leary’s example of protein localization

Answer 3

disrupt cell membrane; use differential centrifugation to separate the parts of the cell; incubate each organelle;part of the cell with Abs specific to the protein you’re trying to detect–> run western blot–> locate protein

Question 4

Describe GFP/RFP method to localize proteins

Answer 4

fuse protein with GFP gene–>when expressed, your protein will give off green fluorescence; fuse another cell with RFP as a cellular location marker (e.g. binds to DNA in nucleus–>localizes nucleus)–> overlay to images; if there is a colur change (GFP/RFP to orange-y colour)–>you have localization and know where your protein is