Experiment #10

Rf

(distance traveled by component) / (distanced traveled by solvent front)

one difference between these two types of polymers (proteins and DNA) in terms of their composition

-proteins are polymers made of amino acids, the group of molecules with amino group and carboxyl group on the same carbon atom -DNA are polymers of nucleotides that are composed of a nitrogenous base, a sugar, and phosphate group

In the paper chromatography experiment, if an amino acid moved 3.6 ± 0.1 cm and the solvent moved 6.0 ± 0.1 cm on the chromatography paper, what is the Rf value of the amino acid with error?

0.60±0.03

Draw the complimentary DNA strand of 5'-T-T-A-C-G-A-G-C-G-3'

3'-AATGCTCGC-5'

Sketch the structure of a micelle and explain how the micelle keeps the casein proteins soluble (how does soap make oil dissolve in water?).

A casein micelle has hydrophobic proteins on the inside, held together by colloidal calcium phosphate. Hydrophilic proteins are on the outside of the micelle, keeping the entire structure suspended in solution

Explain why casein precipitates when acetic acid is added to it.

Acetic acid lowers the pH of the milk, breaking up the micelle by dissolving the colloidal calcium phosphate and bringing casein to its isoelectric point.

Biologically, the casein micelle is considered a calcium phosphate delivery system. Explain.

Calcium phosphate is not soluble at the pH of milk. The casein micelle keeps the calcium phosphate suspended in solution so it can be delivered through the milk.

Casein has an isoelectric point at pH 4.6. What kind of charges will be on the casein in its native environment, that is, in milk (pH about 6.6)?

Casein will be above its isoelectric point in milk. Therefore, both protons will be removed and the overall charge will be negative.

What is the role of calcium phosphate in the casein micelle?

Colloidal calcium phosphate holds together the sub-micelles within the casein micelle.

What is the charge on the sugar-phosphate backbone? How will we exploit this charge, allowing us to isolate the DNA from an onion?

DNA is readily soluble in water due to the negatively charged sugar-phosphate backbone. By adding ethanol, which is less polar than water, DNA becomes less soluble.

What substance is on your hands that will prevent the onion DNA from spooling properly? Why?

DNAase is present on the skin and will cleave DNA.

Why is milk or raw egg used as an antidote in cases of heavy metal ion poisoning?

Heavy metals denature the disulfide bonds in proteins by bonding with the sulfur atoms. Milk and raw egg contain large amounts of protein and therefore complex with the metal and then can be excreted from the body via vomiting

What does the term hydrophobic mean? Hydrophilic?

Hydrophobic: avoid interacting with water; hydrophilic: interact readily with water

List the following in order of increasing polarity IN an aqueous solution: Methanol (CH3OH), Methane (CH4), Water (H2O), Butanol (C4H9OH), Octane (C8H18), Ethanol (C2H5OH)

Octane<Methane<Butanol<Ethanol<Methanol<Water

Biuret is the most common test for proteins. Would a free amino acid, such as proline, give a positive biuret test?

Proline is an amino acid and does not contain a peptide bond. Only proteins give a positive result in the Biuret test.

zwitterion

a molecule or ion having separate positively and negatively charged groups

four different bases

adenine (A), thymine (T), guanine (G), cytosine (C)

Precipitation of casein protein step 2

aggregation of casein protein, isoelectric pH of casein is 4.6

When are proteins least soluble in water?

at their isoelectric pH due to aggregation by electrostatic attraction (therefore more soluble at lower/higher pH)

Precipitation of casein protein step 1

breaking of micelles: calcium phosphate dissolves in acidic solution, free casein protein from micelles

amino acids react with each other through...

condensation to form a long chain polymer-protein

Paper Chromatography: Separating Amino Acids -what is separation based on?

difference in amino acids

Paper Chromatography: Separating Amino Acids -stationary phase

highly polar (paper fiber with water in air)

Paper Chromatography: Separating Amino Acids -mobile phase

less polar (solvent: mixture of water, 1-butanol, and acetic acid)

further the amino acid travels...

less polar they are (in mobile phase with solvent

what do casein proteins exist as

micelles-colloidal particles

Will a less polar species have a large or small Rf value when using paper chromatography to separate components of a mixture? Explain using the terms affinity, mobile phase, and stationary phase.

mobile phase is usually composed of a solvent that is less polar than the stationary phase. Since a less polar species will be attracted to and have a higher affinity for the mobile phase, it will move up the paper more readily. Therefore, Rf value will be large.

Will a more polar species have a large or small Rf value when using paper chromatography to separate components of a mixture? Explain using the terms affinity, mobile phase, and stationary phase.

mobile phase is usually composed of a solvent that is less polar than the stationary phase. Since a more polar species will be attracted to and have a higher affinity for the stationary phase, it will not move very far up the paper. Therefore, the Rf value will be small.

amphiphiles

molecules that contain both a hydrophobic and hydrophilic end

monomer

nucleotide composed of a nitrogenous base, a five-carbon sugar and at least a phosphate group

isoelectronic point

pH at which amino acid is neutral (the zwitterion form is dominant)

Both proteins and DNA are...

polymer molecules

DNA

polynucleotide molecule in cells of living organisms that contains genetic information

Heavy Metal Test

precipitate protein by denaturing protein (change structure thus function by breaking disulfide (S-S) bonds in protein)

Xanthoprotein Test

presence of benzene rings: concentrated HNO3 acid reacts with phenyl ring to give a yellow or orange-colored aromatic nitro compound

Ninhydrin Test

presence of free amino (-NH2) groups: -NH2 forms a purple complex with ninhydrin

Biuret Test

presence of peptide bonds: -NH- in peptide bonds form a purple complex with Cu2+

Sketch the shape of the titration curve for alanine with NaOH. Mark the points where the pKa's of the carboxylic acid and the amino groups can be determined.

refer to graph

micelle

small, nearly spherical structure formed when amphiphiles dissolved in water

Paper Chromatography: Separating Amino Acids -what are polar components more attracted to?

stationary phase, moving along more slowly

what do interactions between the side chains of amino acids in the polymer chain determine?

structure of protein

Explain in your own words the placement of each of the following components in the DNA structure: nucleotide bases, sugar-phosphate backbone, and hydrogen bonds.

sugar-phosphate backbone holds the nucleotide bases in a double helix structure; nucleotide bases are on the inside of the helix interacting through hydrogen bonds

main function of a casein micelle

to carry large amount of highly insoluble calcium phosphate (Ca9(PO4)6) in liquid form

Why do we use ice-cold ethanol to isolate the onion DNA?

to decrease the solubility of DNA.

If a protein contains no tyrosine, could it give a positive xanthoprotein reaction?

xanthoprotein test checks for the presence of a phenyl ring. Since other amino acids have a phenyl ring, it is possible to obtain a positive test without tyrosine being present.