Biochemistry Lecture Exam 3 Flashcards
Carbohydrate
aldehydes or ketones with at least two hydroxyl groups, or substances that yield such compounds on hydrolysis
Monosaccharides
simple sugars, consist of a single polyhydroxy aldehyde or ketone unit
Oligosaccharides
short chains of monosaccharides units joined by glycosidic bonds
Disaccharides
oligosaccharides with two monosaccharide units
Polysaccharides
sugar polymers with 1+ monosaccharides units
Epimers
two sugars that differ only around one carbon atom
Pyranose
six membered ring compounds
Furanose
five membered ring compounds
What is an aldose?
carbonyl group is at an end of the carbon chain (in an ALDEHYDE group)
What is a ketose?
carbonyl group is at any other position but the end (in a KETONE group)
Which isomer (D or L) are most sugars in nature?
D
For glucose, what carbon is the carbonyl carbon?
Carbon 1
For glucose, what carbon is the anomeric carbon?
Carbon 1
What cyclical form does glucose typically take?
Pyranose form is most common
What is an aldonic acid?
form following oxidation of the carbonyl carbon of aldoses
What is an uronic acid?
form following oxidation at carbon 6
What position of the sugar is the reducing end? How can that be used to detect sugar in a solution?
-Reducing End: the end of a disaccharide or polysaccharide chain with a free anomeric carbon
-Produces color change when combined with different sugars (we did this in lab so I think this is right)
What is a homopolysaccharide?
contain only one single monomeric sugar species; serve as storage forms and structural elements
What is a heteropolysaccharide?
contain 2+ kinds of monomers; provide extracellular support
Name two biologically important homopolysaccharides?
starch and glycogen
What are some roles of polysaccharides?
Mainly have structural and energy storage roles
What is the structure of glycogen? What types of bonds occur and what types of sugar is involved?
-Heavily hydrated with many exposed hydroxyl groups that are available to hydrogen bond
-Most stable 3D structure is a helical structure with six residues/turns; more extensively branched and more compact than starch
-made of glucose sugar
What are glycosaminoglycans and what types of modifications are made to the sugars found in GAGs?
-Glycosaminoglycans: heteropolysaccharides found in the ECM; linear polymers composed of repeating disaccharide units; unique to animals and bacteria
-Different groups on the GAG is the modifications (amine group, sulfate added, carboxyl added)
Glycoconjugate
biologically active molecule consisting of an informational carbohydrate joined to a protein or lipid
Proteoglycan
macromolecules of the cell surface or ECM consisting of 1+ sulfated glycosaminoglycan chain(s) joined covalently to a protein or secreted protein; major component of extracellular matrices
Glycoprotein
have one or several oligosaccharides joined covalently to a protein; found on the outer surface of the plasma membrane
Glycolipid
plasma membrane components in which the hydrophilic head groups are oligosaccharides
What features of GAGs cause proteoglycan aggregates to form larger structures in the ECM?
-The 1+ sulfated glycosaminoglycan chain joins covalently to a membrane protein or secreted protein?
What is the difference between N and O linkages in glycoproteins?
-N-linked: an N-glycosyl bond joins the anomeric carbon of a sugar to the amide nitrogen of an Asn residue
-O-linked: a glycoside bond joins the anomeric carbon of a carbohydrate to the -OH of a Ser or Thr residue
What is a lectin and what does it do biologically?
-Lectin: bind carbohydrates with high specificity and with moderate to high affinity
-Functions: cell-cell recognition, signaling, adhesion, intracellular targeting of newly synthesized proteins
Nucleotide
a nucleoside phosphorylated at one of its pentose hydroxyl groups
Nucleoside
a compound consisting of a purine or pyrimidine base covalently linked to a pentose; the molecule without the phosphate group
Gene
a chromosomal segment that codes for a single functional polypeptide chain or RNA molecule
rRNA
components of ribosomes
mRNA
intermediates in protein synthesis
tRNA
adapter molecules that translate the information in mRNA into a specific amino acid sequence
ncRNA
wide variety of functions (Does NOT code for proteins)
Purine
a nitrogeneous heterocyclic base that is a component of nucleotides and nucleic acids; A and G
Pyramidine
a nitrogenous heterocyclic base is a component of nucleotides and nucleic acids; C, T, and U
How do pyramidine and purine structures differ? What nucleotides will base-pair using Watson-Crick pairing?
-Pyrimidines are planar and purines have a slight pucker
-Purines have a 2-ringed structure and pyrimidines only have one
-A base pairs with T (or U in RNA) with 2 H bonds
-G base pairs with C with 3 H bonds
What is a phosphodiester bond? What charge does it have?
-Phosphodiester bond is a covalent bond that joins successive nucleotides of both DNA and RNA between the 5’ phosphate of one nucleotide and the 3’ hydroxyl of the next nucleotide
-Negative Charge
Why is DNA inherently more stable than RNA?
-RNA is rapidly hydrolyzed due to the 2’ hydroxyl groups
What wavelength of light do nucleotides absorb the most? What happens to light absorption when DNA strands denature?
-260 nm is the strongest absorbtion
-More light is absorb when DNA strands are denatured
What type of double helix is found in nature?
-B form
What is the difference between a nucleotide and a nucleoside?
-Nucleotides have 3 components: a nitrogenous base, a pentose, and a 1+ phosphate
-Nucleoside is the molecule without a phosphate group
What are the common classes of RNA and what are their functions?
-rRNA: components of ribosomes
-mRNA: carries genetic information to the ribosome
-tRNA: adapter molecules that translate the information in mRNA into amino acid sequences
-ncRNCA: wide variety of functions; do not code for proteins
What are some structural characteristics of RNA?
single stranded, ribose sugar, 2’ hydroxyl
What is annealing?
process by which 2 strands spontaneously rewind when temperature or pH is returned to its normal range
What type of damage is typically caused by UV light exposure?
pyrimidine dimers
What nucleotide base is most subject to deamination? What does it become?
Cytosine turns into uracil
Where does the chemical energy of a nucleotide come from?
-Hydrolysis of nucleoside triphosphates (ATP)
-Has 3 bonds between different phosphates and the breaking of those bonds creates energy
Genome
the complete haploid genetic complement of an organism
DNA Cloning
selective amplification of a particular gene or DNA segment so that its genetic information may be studied and utilized
Recombinant DNA
composite DNA molecules comprimised of covalently linked segments from 2+ sources
Cloning Vectors
small DNAs campable of autonomous replication
Restriction Enzymes/Endonucleases
recognize and cleave DNA at specific sequences
Ligase
joins the DNA fragments to be cloned to a suitable cloning vector
Plasmid
circular DNA molecule that replicates separately from the host chromosome
Shuttle Vectors
plasmids that can be propagated in cells of 2+ species
What is cloning and how is it used to study/modify a region of DNA?
-DNA cloning: selective amplification of a particular gene or DNA segment so that its genetic information may be studied and utilized
What type of sequences do restriction endonucleases target and how does that lead to the formation of sticky/ blunt ends for cloning?
-Restriction endonucleases target palindromic sequences and make staggered cuts which leads to blunt and sticky ends
What is a plasmid and how can plasmids be used to bring DNA into the cell?
Plasmids are circular DNA molecules that replicate separately from the host chromosome that usually have a symbiotic role in the cell
What is the difference between selectable marker and screenable marker?
-Selectable Marker: either permits the growth of a cell (positive selection) or kills the cell (negative selection) under defined conditions
-Screenable Marker: gene encoding a protein that causes the cell to produce a colored or fluorescent molecule
What are some strengths and weaknesses of bacterial expression systems?
-Strengths: regulatory sequences are well understood, can express high levels of cloned proteins, easy to store and grow, efficient methods for transforming and extracting DNA, can be grown in huge amounts
-Weaknesses: some heterologous proteins do not fold correctly, proteins may not undergo necessary posttranslational modifications or proteolytic cleavage, some gene sequences can be difficult to express
What are some strengths and weaknesses of yeast expression systems?
-Strengths: well-understood eukaryotic organism, expression of eukaryotic genes can be more efficient, proteins may be folded and modifies more accurately
-Weaknesses: heterologous proteins may not fold properly, yeast may lack the enzymes needed to modify the proteins to their active forms, certain features of the gene sequence may hinder expression of a protein
What methods can be used to change a nucleotide sequence in order to change an amino acid sequence?
Oligonucleotide-Directed Mutagenesis or Site-Directed Mutagenesis
Fusion protein
product of a ligated gene containing parts of two different genes
Tag
peptide or protein that binds to a sample, stable ligand with high affinity and specificity
RT-PCR
uses reverse transcriptase to generate a DNA strand from an RNA template, followed by the standard PCR protocols using DNA polymerase
qPCR
is used to estimate relative copy numbers of particular sequences in a sample
cDNA
double-stranded DNA fragments formed from mRNA templates (relies on reverse transcriptase)
Transcriptome
the entire complement of transcribed RNAs present at a given moment in the cell
Proteome
the entire complement of proteins present at a given moment in a cell
How can tags be used to specifically purigy proteins or identify their location in a cell?
-fuse to gene encoding target protein and permits purification by affinity chromatography
-help to provide good yield and high purity and may affect the properties of attached proteins
How are PCR and qPCR similar and how do they differ?
-Both amplify DNA using primers and DNA polymerase
-qPCR measures fluorescence in real time and gives a quantitative measure of the results
How does immunoprecipitation work?
-Immunipreciption: process of precipitating a fusion protein (containing the gene of interest and a gene for an epitope tag) by antibodies to the epitope
-proteins that bind to the tagged protein will also precipitate
How does tandem affinity purification work?
-Tandem Affinity Purification (TAP) tags: two consecutive tags that are fused to a target protein to enhance the selectivity of immunopreciption
-First tag (protein A) binds mammalian IgG
-Second tag (calmodulin-binding peptide) binds calmodulin
What is a yeast two-hybrid experiment? What can you learn from it?
-technique that relies on the properties of the GaI4 protein
-two domains of GaI4p must be brought together to function correctly
-probes molecular interactions in vivo
How does the CRISPR-Cas9 system work?
-The CRISPR/Cas Complex binds and the Cas9 two separate domains cleave opposite strands of DNA
-The sgRNA guide sequence can be altered to target any genomic sequence
-required to pair with the target DNA sequence and to activate the nuclease domains
-cells in which genes required to survive the treatment are inactivated or activated by teh CRISPR/Cas9 varient will die or thrive
