PROTEIN_ELECTROPHORESIS Flashcards
In protein electrophoresis, proteins can be separated from one another by _____. In the example at left, each dark line is a _____ – a bunch of identical molecules of one kind of protein. Each vertical column of bands is a
_____. Within one lane, all the proteins started together at the ____, but the _____ proteins moved faster and ended up closer to the bottom.
size band lane top smaller
In protein electrophoresis, you force
proteins to migrate through a gel. The gel
can be any material that is solid, but has
pore spaces large enough for the proteins to fit through. If the pores are a tight fit for the proteins, the larger proteins will move
_____, while the smaller proteins will be
_____. You make the molecules move through the gel by applying an ______ across the gel. _______molecules will move toward the negative pole, and _______ molecules will move toward the positively charged pole. ________ won’t move at all.
slowly free to move faster electric field Positively charged negatively charged Uncharged molecules
A protein’s rate of movement through the
gel will be controlled by:
Size Charge Shape Pore size Voltage
A protein’s rate of movement through the gel will be controlled by: • Size. \_\_\_\_\_ molecules move more slowly, because they don’t fit through the pores easily.
• Charge. \_\_\_\_\_\_\_\_\_ of charge influence how fast and which direction a molecule moves. Some proteins are positively charged and some are negative; in most cases, the charge will depend on pH and other aspects of the solution.
• Shape. A protein that is ________
will seem smaller, and move through
the gel faster, than a protein that is
________.
• Pore size. \_\_\_\_\_\_\_=faster movement. \_\_\_\_\_\_\_\_\_ may allow more precise separations. Pore size varies from gel to gel, but within a single gel, each \_\_\_\_ has the same pore size.
• Voltage.The more voltage you apply
across the gel, the _____ things will
move. Within a single gel, voltage is
____.
Bigger Strength and polarity tightly folded; loosely folded Bigger pores; Smaller pores; lane faster; constant
It is a way of doing electrophoresis which ensures that a band’s rate of migration is determined only by size
SDS-PAGE
SDS stands for ________
Sodium Dodecyl Sulfate
SDS is a detergent, and it accomplishes several critical things for electrophoresis:
• SDS helps proteins \_\_\_\_\_\_so you can run them on the gel (not all proteins are soluble in plain water). • SDS \_\_\_\_\_\_, or unfolds, proteins. This means that shape will not influence rate of migration in the gel. • SDS sticks to proteins and makes them \_\_\_\_\_\_. Since SDS sticks all over the protein, each protein ends up with the same density of charge.
dissolve
denatures
negatively charged
PAGE stands for ___________. You should probably remember this. ________ is used because it creates strong gels with a
predictable pore size.
Polyacrylamide Gel Electrophoresis
Polyacrylamide
In SDS-PAGE, all the proteins have the same
________ and the same ________. In a single gel, the pore size and the voltage are ______. Therefore, in SDS-PAGE, a protein’s rate of migration through the gel is determined solely by ______.
charge density
unfolded shape
constant
size
_______ is the most commonly used method of protein electrophoresis.
SDS-PAGE
THE PROTEIN SAMPLES FOR THIS LAB
Almost any ______ tissue can be used for
SDS-PAGE. We’ll use ____, because it’s
mostly protein and easy to grind up
biological
fish
GEL STAINING
You need to stain the proteins before you
can see them. In today’s lab, you’ll use a
stain called _____. This chemical was
originally developed as a dye for wool, but it
happens to stain most other proteins pretty
well, too. After you run your gel, you’ll soak
it in a ““_______ so you can see
the bands.
Coomassie
Coomassie solution
________ is similar to electrophoresis, but the key difference is that in chromatography the proteins are pulled through a gel or similar matrix by a moving solvent, rather than an electric field.
Chromatography
