Brainscape's Knowledge GenomeTM

Browse over 1 million classes created by top students, professors, publishers, and experts.


See full index

BMS > Tools of Protein Biochemistry > Flashcards

Tools of Protein Biochemistry Flashcards

1
Q

What are two major types of electrophoresis

A
  1. ) Native-PAGE (non-denaturing)

2. ) SDS-PAGE (denaturing)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
2
Q

Separates molecules according to their charge, shape, and size

A

Electrophoresis

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
3
Q

How many proteins are expressed in our bodies?

A

20,000

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
4
Q

Buffer pH and intrinsic chemical properties of macromolecules (DNA, RNA, and proteins) determine their

A

Net charge

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
5
Q

Electrophoresis requires a gel for

A

Support and Molecular sieving

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
6
Q

What does PAGE mean in SDS-PAGE?

A

Polyacrylimide gel electrophoresis

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
7
Q

Electrophoresis where the protein retains it’s activity throughout the process

-Non-denaturing

A

Native-PAGE

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
8
Q

Polypeptides retain their higher-order structure and often retain enzymatic activity and interaction with other polypeptides in

A

Native PAGE

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
9
Q

In native PAGE, the migration of proteins depends on

A

Size, shape, and native charge

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
10
Q

Single amino acid substitution of Glutamate 6 with valine

-causes conformational change that affects hemoglobins ability to bind oxygen

A

Sickle Cell Disease

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
11
Q

Hemoglobin A is a tetramer composed of what 4 subunits?

A

2α and 2β

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
12
Q

What significance does the fact that a glutimate (negative charge) is substituted for a valine (neutral charge) in people with sickle cell?

A

HbA and Hb-S can be separated at physiological pH because HbA is negatively charged

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
13
Q

Which Hb migrates faster during electrophoresis at pH 7?

A

HbA (it is negatively charged)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
14
Q

Which Hb migrates faster during electrophoresis at pH 4?

A

HbA and Hb-S have the same charge and thus can not be separated

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
15
Q

Electrophoresis method to separate proteins baed on molecular weight

A

SDS-Page

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
16
Q

Prior to SDS-PAGE, proteins are treated with which two things?

A
  1. ) Reducing agent to break disulfide bonds (β -mercapthanol)
  2. ) SDS
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
17
Q

An ionic detergent that denatures secondary and non-disulfide-linked tertiary structures

-Binds to protein backbone and confers a negative charge proportional to mass

A

Sodium dodecylsulfate (SDS)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
18
Q

What fragments moves faster in SDS-PAGE?

A

Small fragments

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
19
Q

The movement in SDS-PAGE is not linear, but rather

A

Logarithmic

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
20
Q

Cyclists, distance runners and crosscountry skiers have been utilizing recombinant human erythropoietin (RhEPO) to improve their endurance performance. How can we detect Erythropoietin?

A

SDS-PAGE

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
21
Q

Induces hemoglobin production which allows more oxygen to be carried in the blood and improves endurance

A

Erythropoietin

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
22
Q

Erythropoietin was created for people with anemia or people undergoing chemo to help increase

A

Hemoglobin production

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
23
Q

Erythropoietin (EPO) is a hormone produced by the kidney that promotes the formation of

A

Red blood cells by bone marrow

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
24
Q

Blood does not have many circulating proteins in it, but in the cell their are many. How can we isolate a specific protein of interest?

A

Western Blot

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
25
Q

In all types of blotting,we make our molecules negatively charged so that we are separating exclusively by

A

Size

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
26
Q

How do we generate antibodies that bind our target protein in a western blot?

A

Inject a rabbit with the antigen (target protein). Take serum from the rabbit. The supernatant will contain the desired antibodies

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
27
Q

Portion of a molecule to which an antibody binds

A

Epitope

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
28
Q

The antigen binding site of an antibody

A

Paratope

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
29
Q

Epitopes can be composed of

A

Sugars, lipids, or amino acids

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
30
Q

Antibody with a bound enzyme that can catalyze

conversion of a colorless molecule to a colored one

A

Indicator

31
Q

What are two common indicators?

A

Alkaline phosphatase and Horseradish peroxidase

32
Q

Summarize a western blot

A

The antibodies specific to the protein of interest are generated by injecting a rabbit with the antigen (protein of interest) and taking blood serum from the rabbit. Next, gel electrophoresis is performed on the sample containing our target protein. Upon completion, the wells are transferred from the gel to a nitrocellulose sheet. The antibodies we generated are added to the sheet, at which point they bind the target protein. All other antibodies are washed way and tagged antibody is added (indicator), which allows us to isolate the desired protein.

33
Q

Combine electrophoresis and immunological techniques

A

Western Blots

34
Q

First thing you do when screening for a disease. A faster, more sensitive method than western blotting to detect antibodies or antigens in a patient

A

Enzyme-Linked Immunoabsorbant Assay (ELISA)

35
Q

Antibodies developed by the human body against AIDS can be detected by

A

ELISA

36
Q

Troponin, released after a myocardial infarction, can be detected by

A

ELISA

37
Q

Which is more specific, Western Blotting, or ELISA?

A

Western Blotting

38
Q

Explain how ELISA works

A

You have a well, to which you can link an antigen or an antibody, depending on which one you are testing for. Next, add an enzyme antibody, which will bind to the antigen and allow for detection.

39
Q

Tagged antibodies bind directly to the antigen in the well and allow for detection in?

A

Direct ELISA

40
Q

A primary antibody binds directly to the antigen in the well. Then, tagged secondary antibodies bind to the primary antibody and allow for detection in?

A

Indirect ELISA

41
Q

How is the antigen actually detected in a western blot or ELISA?

A

A substrate is added that binds the enzyme attached to the antibody and produces either a fluorescent or colored product

42
Q

ELISA is more sensitive than a western blot because?

A

The well is hooked up to a spectrophotometric detector. Where as in a western, we have to observe the color change with our eyes (requires a larger sample)

43
Q

What is the gold standard for HIV testing?

A

Perform an ELISA, if it’ negative than the patient is negative. If it is positive, perform a Western blot for confirmation. The blot results will provide the official diagnosis

44
Q

The time between HIV infection and when antibodies to HIV are produced

A

Window Period (4-6 weeks)

45
Q

What indicates a positive HIV Western Blot?

A

You need one band to be present in the region of gp 160 or gp 120, and one band present in the region of p31 or p24

46
Q

When bands are present but the pattern does not meet the criteria for positivity, the result is

A

Indeterminate

47
Q

What are some causes of an indeterminate HIV western blot?

A

Recent infection, advanced HIV, certain strands of HIV, etc.

48
Q

What is the procedure following an indeterminte result?

A

Retest in > 6 weeks

49
Q

What does the Western blot HIV test actually measure?

A

The antibody to HIV

50
Q

What are the antibodies to HIV bound to?

A

Antigen to HIV

51
Q

Inadequate blood supply to an organ or part of the body

-Cause of myocardial infarction

A

Ischemia

52
Q

Serve as useful diagnostic markers of acute ischemia

A

Troponin, Myoglobin, and Creatine Kinase

53
Q

Cardiac cells rupture and release proteins into the blood. As a consequence, the levels of Troponin T increase, which indicates

A

Myocardial infarction

54
Q

The contractile regulating protein of striated muscle

A

Troponin

55
Q

The cardiac isoforms of troponin T and I increase in serum after

A

Acute Myocardial Infarction

56
Q

Troponin is highly specific for myocardial infarction because it is not elevated in any other trauma, heavy exercise, or renal failure. Also, it remains in the blood for up to

A

Four days following the event

57
Q

What is the difference between HIV ELISA and Troponin ELISA?

A

Troponin ELISA uses an immobilized antibody in the well. Where as HIV used an immobilized antigen

58
Q

Describe how co-immunoprecipitation works

A

We bind beads to an antibody specific to our target protein and add it to our sample. The antibody binds the target protein.The increased mass from the beads causes the complex to precipitate out. If our target protein is complexed with another protein, that protein will also precipitate out. We can separate the two by SDS page and then perform a western blot

59
Q

Allows the proliferation of damaged cells, which can lead to tumor development

A

Loss of p53

60
Q

p53 tumore suppressor is frequently disabled by

A

Mutations

61
Q

Wild type in 50% of human cancer

A

p53 gene

62
Q

In response to cellular stress, promotes p53 ubiquination and degredation by the proteasome

-An E3 ubiquitin ligase

A

MDM2 (HDM2)

63
Q

Targets p53 for degredation to disable the p53

A

Overexpression of MDM2 in human cancer

64
Q

How did scientists show that MDM2 and p53 interact?

A

Immunoprecipitation of MDM2 which showed coprecipitation of p53

65
Q

Describe immunoaffinity Chromatography

A

We add antibodies specific to our target protein to a column. We then add a cell extract to the column. As the cell extract moves through the column, the target protein will bind the antibodies and everything else will be eluted. Then we prepare a solution to elute our target protein, which we can then load to the gel and quantify

66
Q

Purifying proteins from cell extracts

A

Affinity Purification

67
Q

Can immunoaffinity chromatography be used for interaction proteins similar to how co-immunoprecipitation worked?

A

Yes

68
Q

Uses primary antibodies to target and label specific proteins in tissue and then secondary antibodies with a conjugated enzyme to bind the primary antibodies

-Key is that it is in the tissue

A

Immunohistochemistry

69
Q

A breast cancer that tests positive for a protein called human epidermal growth factor receptor 2 (HER2), which promotes the growth of cancer cells

A

HER2-positive breast cancer

70
Q

In about 1 out of every 5 breast cancers, there is a gene mutation that results in the roduction of excess

A

HER2

71
Q

Tend to be more aggresive than other types of breast cancer

A

HER2-positive breast cancers

72
Q

Added to molecules that an investigator wants to visualize when we can’t develop an antibody for our protein of interest

A

Epitope tag

73
Q

How does epitope tagging work?

A

We make and add a tag to the DNA sequence of the desired molecule, which results in the expressed protein being made with the tag in its amino acd sequence. We can then generate an antibody specific to the tag and detect with a blot

BMS (62 decks)

Brainscape helps you reach your goals faster, through stronger study habits.
© 2024 Bold Learning Solutions. Terms and Conditions