Cell Biology- Protein Synthesis

Beta bends are normally composed of...

4 amino acids, one of which is often proline (this causes a kink in the polypeptide chain)

Degeneracy (genetic code)

Degeneracy: also called redundancy = a given amino acid may have more than one triplet coding for it (Arg is coded by six different codons)

Function of Enzymes and peptide hormones

Direct and regulate metabolism

E site

E site: occupied by the empty tRNA as it isabout to exit the ribosome

missense mutation

Missense mutation: The codon containing the changed base may code for a different amino acid (UCA → Ser; CCA → Pro)

What types of bonds make up the primary structure?

peptide bonds- between C-atom of carboxyl group of 1 AA and N-atom of amino group of another AA

Are globular proteins soluble in water?

yes! its readily soluble.

Where are ribosomes found?

"free" in the cytosol or in close association with the endoplasmic reticulum(rER)

AA's with uncharged polar side chains have a __ net charge at physiologic pH. a. Zero b. Positive C. Negative

0

STEPS IN PROTEIN SYNTHESIS

1. Activation 2. Initiation 3. Elongation 4. Termination 5. Folding and posttranslational modifications

Name some types of structure of proteins

1. Fibrous proteins 2. Globular proteins 3. Intermediate proteins

Name some examples of Composition Proteins

1. Simple proteins 2. Glycoproteins 3. Nucleoproteins 4. Chromoproteins 5. Lipoproteins 6. Flavoproteins 7. Metalloproteins

Name some examples of functional proteins

1. Structural proteins 2. Enzymes 3. Hormones 4. Respiratory pigments 5. Transport proteins 6. Contractile proteins 7. Storage proteins 8. Immunoglobulins 9. Toxins

Selenocysteine

21st proteinogenic AA, not coded directly by genetic code

Peptides are different from proteins as they have ___ or fewer amino acids

50

A site

A site: binds an incoming aminoacyl-tRNAaccording to the codon occupying the site

Activation of protein synthesis (mention what happens and where it occurs)

AAs are activated by special aminoacyl-tRNA synthetases in the cytosol, AA attached to tRNA

What happens to all the excess AA's? (amino acid catabolism) (Nitrogen metabolism)

All amino acids in excess of biosynthetic needs are rapidly degraded (transamination following oxidative deamination in liver and kidney) → into ammonia (NH3) and corresponding α-keto acids, the carbon skeletons of AA In the second phase of amino acid catabolism, the carbon skeletons of the α-keto acids are converted to common intermediates of energy-producing metabolic pathways, which can be metabolized to CO2 and H2O, glucose, fatty acids, or ketone bodies by the central pathways of metabolism (gluconeogenesis, glycolysis, TCA cycle)

Where do peptides bond?

Amino acids are linked by peptide bonds between theα-carboxyl group of one amino acid and the α-amino group of another

When there is a lot of misfolded proteins, what sort of diseases can happen? (fyi)

Amyloid diseases (FYI) • Abnormal conformation of fibrillary protein assemblies of β-pleated sheets called amyloids • Implicated in degenerative diseases such as Alzheimer, Parkinson, and Huntington Prion diseases (FYI) - Infectious protein is an altered version of a normal prion protein, it forms insoluble aggregates of fibrils - E.g. Creutzfeldt-Jakob disease in humans, scrapie in sheep, bovine spongiform encephalopathy in cattle ('Mad Cow' disease)

Proteins as Enzymes

Biological catalysts, reduce activation energy of reactants and speed up reactions Most of them are globular, conjugated proteins, Examples: DNA Polymerase, Lipase

contractile proteins function

Can contract muscles with the expense of energy from ATP Examples: Actin, Myosin

WHAT SORTS OF DISEASES COULD HAPPEN IF THERE WAS A CHANGE IN CRITICAL AMINO ACIDS IN A PROTEIN?

Changes in amino acids (→ mutations in the DNA)can lead to a non-functional protein or to a misfolded one • Examples: sickle cell anemia, Alzheimer disease,transport defects, enzyme deficiencies

Regard conjugated proteins, what classifies the different types?

Classification based on the nature of the prosthetic groups (p.g.): a) Phosphoproteins: p.g. is phosphoric acid (e.g. Casein/milk, Vitellin/egg) b) Glycoproteins: p.g. is carbohydrate (most membrane proteins) c) Nucleoproteins: p.g. is nucleic acid (proteins in chromosomes, ribosomes) d) Chromoproteins: p.g. is pigment or chrome (Haemoglobin, Phytochrome) e) Lipoproteins: p.g. is lipid (some membrane proteins, chylomicrons) f) Flavoproteins: p.g. is FAD- Flavin Adenine Dinucleotide (proteins of electron transportsystem) g) Metalloproteins: p.g. is metal ions (Nitrate Reductase)

Proteins as Respiratory pigments, give an example

Coloured, conjugated proteins with a pigment (chrome) as their p.g. Examples: Haemoglobin, Myoglobin

conjugated proteins structure

Complex, contain one or more non-amino acid components usually globular These non protein parts are called prosthetic groups, essential for biological function

Structural proteins are in

Components of connective tissue, bone, tendons, skin, feathers, nail, hair, horn Mostly fibrous proteins, insoluble in water, Examples: Collagen, Keratin, Elastin

Simple proteins composition

Composed of only amino acids, relatively simple structure Examples: Myosin, Collagen, Keratin, Insulin

1. Describe the flow of information according to the central dogma of molecular biology.

DNA → RNA → Protein Transcription → Splicing →Translation

What types of bonds stabilize the tertiary structure?

Disulfide bonds Hydrophobic interactions Hydrogen bonds Ionic interactions

Protein translation

Each tRNA has an attachment site for a specific amino acid at its 3'-end and an anticodon that pairs with a specific codon on the mRNA (adapter combining amino acids with codons)

Causes of mutations (name 3)

Errors during replication of DNA: esp. during meiosis, when cells divide to produce gametes Damage to DNA: via Radiation, Carcinogens and other toxins, Viruses and bacteria Errors during repair! Mutations can cause negative or positive changes, and are important for evolution, but also cause pathology

The polypeptide backbone does assume a random three-dimensional structure. T/F

False, It generally forms regular arrangements of amino acids located near each other in linear sequence (secondary structures)

Function of Immunoglobulins

Fight infectious bacteria and viruses

Function of Structural proteins (collagen, keratins)

Form tissues and bones, acting like steel cables in reinforced concrete

what?

Here's a meme break, you got this!

Describe the overall formula of an alpha helix. (Be sure to also mention how many AA's per turn , type of bond, the shape of the structure)

Hydrogen bonds between carboxy- and amino groups of amino acids (distance of4 AA) → spiral structure(example keratin)

Are fibrous proteins soluble in water?

Insoluble in water

When folding proteins, what sort of interactions determine how long the pp (polypeptide) chain folds and the shape (3D shape)?

Interactions between the side chains of amino acids determine how a long polypeptide chain folds into the three dimensional shape of the functional protein

Intermediate proteins structure

Intermediate shape more or less linear

Where does post-translational modification occur on the AA?

It can occur on the amino acid side chains, or at the C- or N- termini

Cysteine, why is the amino acid special and different from the other AA's?

It has an SH bond that is slightly polarized BUT BUT BUT its a non-polarized molecule

In post-translational modifications of proteins involves what sort of modifications?

It involves the formation of covalent and generally enzymatic modifications of proteins following protein biosynthesis

Why are peptide bonds resistant to conditions that denature proteins such as heating and high concentrations of urea?

It is a strong covalent bond

Describe the structure of fibrous proteins

Linear in shape The secondary structure is functionally most important Usually do not have a (complex) tertiary structure Tough and strong Long parallel polypeptide chains cross-linked at regular intervals

Why does denaturing a protein not denature the primary polypeptide structure?

NO hydrolysis of peptide bonds, primary structure (generally) remains intact

Are polypeptides ever branched? Y/N

No! Its long, continuous, and unbranched

7. What are the four general groups of amino acids, what determines the biochemical behavior of an amino acid?

Nonpolar side chains Uncharged Polar side chains Polar Acidic side chains Polar Basic side chains The side chains are what determine the function of the AA

Nonsense mutation

Nonsense mutation: The codon containing the changed base may become a termination codon (UCA → Ser; UAA → Stop and the protein will be truncated)

How are non-standard amino acids (the non basic 20 AA) synthesized?

Nonstandard amino acids are synthesized by chemical modifications of standard amino acids

How does the body get proteins? (3 ways)

Obtained from the diet (9 AA are essential) Synthesized de novo Released through the degradation of body protein

Where does protein folding occur and how long does it take?

Occurs in the cell in seconds to minutes

P site

P site: is occupied by peptidyl-tRNA whichcarries the chain of amino acids that hasalready been synthesized

What sort of functions do the fibrous proteins do for the cell?

Perform structural functions in the cell

Function of Contractile proteins in muscle (actin and myosin)

Permit movement

22. Name some differences between prokaryotes and eukaryotes with regard to transcription and translation

Prokaryotes → 50S and 30S subunits = 70S R

Proline, why is the amino acid special and different from the other AA's?

Proline differs from other nonpolar amino acids in that its side chain and alpha-amino nitrogen form a rigid, five-membered ring structure It has a secondary (rather than primary) amino group (imino group)

What is a good way to break peptide bonds? (with out enzymes)

Prolonged exposure to strong acid or base at elevated temperature is necessary to break these bonds nonenzymatically

Which structures get denatured during protein denaturation?

Protein denaturation results in the unfolding and disorganization of a protein's secondary and tertiary structures

Hormones give some examples

Proteinaceous hormones such as Insulin, Glucagon, Gastrin, Cholecystokinin/CCK

Ribosomes consist of two subunits. What are they?

Ribosomes consist of two subunits (60Sand 40S in eukaryotes = 80S Ribosome,50/30 S = 70S Ribosome in prokaryotes)

Hydroxylation of tryptophan yields...

Serotonin

21. What is sickle cell anemia, what is its cause, and some effects for sufferers?

Sickle cell anemia is caused by a single nucleotide substitution in the gene for beta-globin missense mutation changes Glufor Val at position 6 The symptoms are characterized by episodes of pain, chronic hemolytic anemia, infections

Name the 3 types of mutations

Silent mutation Missense mutation Nonsense mutation

5. Explain the basic, general shape of an amino acid

So! There is not really a "basic general shape" mentioned. I think she wants us to know the general formula of amino acids and that it is linear an amino group an alpha carbon a side chain and a carboxyl group

globular proteins structure

Spherical/globular shape Polypeptide chain tightly folded into spherical shapes Tertiary structure is functionally most important Physically softer

alpha helix- describe structure

Spiral structure, a tightly packed, coiled polypeptide chain core Side chains of component amino acids extend outward to avoid interfering sterically with each other

What stabilizes the alpha helix structure? (be really specific)

Stabilized by extensive hydrogen bonding between the peptide-bond carbonyloxygens and amide hydrogens

Steps of Elonggation (protein synthesis)

Step 1: Binding of the aminoacyl-tRNA to the A-site of the ribosome Step 2: Generation of the peptide bond in P-site Step 3: Movement of the ribosome along the mRNA then translocates the tRNA from A-site to P-site, ‚empty' tRNA dissociate from E-site

storage proteins function

Store metal ions and... amino acids? found in seeds and pulses, egg and milk Examples: Ferritin (iron), Casein, Ovoalbumin, Gluten

Do subunits work together or independently of each other? (Not a Yes or No answer)

Subunits may function independently of each other, or may work cooperatively, such as in hemoglobin Hemoglobin, binding of oxygen to one subunit increases the affinity of other subunits for oxygen

What makes tertiary structure proteins different from secondary ones?

Tertiary is both the folding of the domains and the final arrangement of the domains in the polypeptide. Secondary forms regular arrangements of amino acids located near each other in linear sequence

Silent mutation

The codon containing the changed base may code for the same amino acid (UCA → Ser; UCU → Ser)

POST-TRANSCRIPTIONAL MODIFICATIONS

The process in eukaryotic cells where primary transcript RNA is converted to mature RNA It is ESSENTIAL for the correct translation of eukaryote genomes

What 3 steps are in the post transcriptional modifications?

The process involves three major steps: 1. Addition of a 5' cap 2. Addition of a 3' polyadenylation tail 3. Splicing (removal of introns (non-coding sequences) and joining of exons (coding sequences important for translation)

What makes an AA unique from other AA?

The side chain (R) R groups can be polar, nonpolar, negatively and positively charged

Why are non-polar AA's side chains considered to be "oily" or "lipid like"?

The side chains promote hydrophobic interactions!

What will happen to a protein when its in an aqueous solution?

The side chains tend to cluster together in the interior protein (hydrophobic effect)

What is the "unique" geometry of proline used for? What is the downfall to this?

The unique geometry of proline aids with the formation of the fibrous structure of i.e. collagen, but interrupts the α-helices in globular proteins

Function of Transport proteins (albumins, hemoglobin)

Transport essential molecules through the bloodstream

What is the fate of newly synthesized proteins? (name 5)

Transport into the Endoplasmic Reticulum→ cisternae Protein folding Transport from the ER to the Golgi apparatus (in vesicles) Glycosylation of newly synthesized proteins takes place in the Golgi apparatus Transport to other organelles

Transport proteins function

Transport materials in cells and form channels, Example: Serum Albumin

2. List the different functions of membrane proteins (I don't think she expects you to know all of these, just at least be able to recognize the limitations of the protein membranes)

Transport proteins Channels Enzymes Signal proteins Hormone receptors Structure proteins Second messengers

Most α-amino acids are found in proteins (proteinogenic) but two of them are not. Which ones are they and how are they synthesized?

Triiodothyronine and thyroxine → thyroid hormones, synthesized from tyrosine slightly changed around AA

Can denatured proteins be renatured?

Under ideal conditions, denaturation may be reversible, so the protein refolds once the denaturing agent is removed Mostly proteins, once denatured, remain permanently disordered (insoluble, precipitate)

1. Regard the Central dogma, what does it mean for proteins?

We get proteins through translation! Proteins are made from information of our genes.

Proteins are the product of our genes. Y/N?

Yes! Think Central dogma!

What determines the role of amino acids in the protein?

a distinctive side chain (R-group)bonded to the α-carbon, this side chain determines the role of the amino acid in the protein

Specificity (genetic code and regard aa)

a particular codon always codes for the same amino acid

@ Physiologic pH the side chains (of AA's with POLAR ACIDIC side chains) are ____ and ____. a. fully ionized, and neutrally charged b. fully ionized, positively charged c. fully de-ionized, negatively charged d. fully ionized, and positively charged e. fully ionized, negatively charged

a. fully ionized, negatively charged

Phosphorylation

addition of a phosphoryl group

Elongation of protein synthesis (also mention what sort of energy is required)

addition of amino acids to the carboxyl end of the growing peptide chain. The ribosome moves from 5'-end to the 3'-end of the mRNA, GTP is required.

What is the most common protein structure?

alpha helix

Are amino acids stored in the body?

amino acids are not stored by the body, no protein exists just for storage of amino acids

What all makes up an amino acid?

an amino group, an acid (carboxyl group), and a distinctive side chain (R-group)bonded to the α-carbon

QUATERNARY STRUCTURE

an arrangement of many polypeptide subunits

2 types of beta sheets

antiparallel and parallel

What are rER-ribosomes responsible for?

are responsible for SYNTHESIZING PROTEINS that are to be EXPORTED from the cell or to be placed in cell membranes (plasma, ER, lysosome membranes)

Initiation of protein synthesis (Mention what happens and where it occurs)

assembly of components of the translation system before peptide bond formation occurs, the formation of ribosome complex (SSU, LSU, mRNA)

Glycosylation

attachment of a glycan (carbohydrate, polysaccharide)

@ Physiologic pH the side chains (of AA's with POLAR BASIC side chains) are ____ and ____. a. fully ionized, and neutrally charged b. fully ionized, positively charged c. fully de-ionized, negatively charged d. fully ionized, and positively charged e. fully ionized, negatively charged (Dr. Freeman says questions like these will definitely be in the exam!)

b. fully ionized, positively charged

which type of beta sheet protein structure are common in nature?

both

Methylation of lysine yields...

carnitine

Polymeric

composed of many repeating monomers (ex: proteins)

Most enzymes are___ proteins

conjugated and or globular

What is the shortest peptide possible?

dipeptides

What happens to free ammonia?

excreted in urine, but most is used in synthesis of urea (CO(NH2)2)→ Urea cycle

Keratins

fibrous protein component of hair and skin, rigidity determined by number of disulfide bonds between constituent polypeptide chains

What sort of things can globular proteins form?

form enzymes, antibodies, and some hormones Examples: Insulin, Haemoglobin, Albumin, DNA Polymerase, and RNA Polymerase

How are Peptide (amide) bonds formed?

formed when carboxyl of one amino acid reacts with the amino group of another

what are domains, in regards to protein?

fundamental functional and three-dimensional structural unitsof polypeptides

Describe Myoglobin structure

globular, flexible molecule, but also highly α-helical

What bonds helps build the pleated structure (beta protein)?

h bonds

What sort of things can denature a protein?

heat, organic solvents, strong acids or bases, detergents, heavy metal ions such as lead

Decarboxylation of histidine yields...

histamine

What types of bonds stabilize the secondary structure?

hydrogen bonds, weak bonds

What stabilizes the quaternary structure protein?

hydrogen-bonds and also by hydrophobic and hydrophilic interactions between AA-R-groups

Under what conditions do condensation reactions occur?

is not a spontaneous reaction, but depends on ATP-dependent activation by attachment of AA to a tRNA (by an enzyme) leaves water!

Universality (genetic code)

it is conserved from very early stages of evolution

What is the common structure shared by all proteins?

linear polymers of amino acid

where does protein metabolism occur?

liver

Acetylation and methylation of serotonin yields...

melatonin

Is protein folding a spontaneous process? (not a yes or no answer, will need to explain answer)

not generally a spontaneous process, but it requires facilitation from a specialized group of proteins: 'Molecular chaperones' or 'Heat shock proteins (Hsp)

Organelles involved in protein synthesis

nucleus, ribosomes, & Golgi apparatus

Termination of protein synthesis (mention when it occurs and how much energy is needed for translation)

occurs when one of the three termination codons moves into the A site, at least 4 high-energy phosphate equivalents (from ATP and GTP) are needed for each peptide bond to guarantee fidelity of translation

Throwback to Block 1: How many amino acids are in oligo peptides and in poly peptides?

oligo: 2-20 AA poly: 21-50 AA

Where is Oxytocin produced?

peptide hormone produced in the hypothalamus (uterine contractions and milk secretion)

Glucagon

peptide hormone, produced by α-cells of the pancreas

CCK (cholecystokinin)

peptide hormone, stimulates pancreas and liver secretion

Gastrin

peptide hormone, stomach hormone that stimulates the secretion of gastric glands

Folding and posttranslational modifications

polypeptides fold into active, 3-dimensional forms; many are processed further

Glycosylation is a... post-translational mod post-transcriptional mod

post transcriptional mod

Phosphorylation is a ... post-translational mod post-transcriptional mod

post transcriptional mod

Atrial Natriuretic Peptide (ANP)

produced in the heart (atria), essential for regulation of blood volume and pressure

antidiuretic hormone (ADH) What is it and where is it produced?

produced in the hypothalamus and essential for the maintenance of water balance (ur- urine related)

AA's with polar acidic side chains are ___ donors. a. electron donors b. Proton donors c. Blood donors d. R group donors

proton donors

AA's with polar basic side chains accept___. a. protons b. electrons

protons

Beta bends, what do they do?

reverse the direction of a polypeptide chain, helping it form a compact, globular shape this makes it crucial for protein structure and function

What sort of solubility does intermediate protein have in the water?

soluble in water eg. blood clotting proteins

What does cytosolic ribosomes do?

synthesize cytosolic proteins or those intended for the nucleus, mitochondria or peroxisomes

Proteinogenic

the ability of certain (20) amino acids to be integrated into proteins

Nonoverlapping and commaless (genetic code)

the code is read from a fixed starting point as a continuous sequence of bases without any punctuation between codons

Creatine

tripeptide involved in energy production in muscle and cardiac cells

Melanin

tripeptide, a pigment occurring in several tissues, synthesized from tyrosine via DOPA

How many polypeptides/polypeptide chains does it take to make a pleated structure?

two or more separate polypeptide chains or segments of polypeptides

3. Describe the basic structure of a protein

unbranched long chains of polypeptides

What normally happens to misfolded proteins?

usually tagged and degraded in the cell

Chaperones

• Chaperones are heat-stable proteins • Also called 'heat shock proteins'(Hsp) • High activity at high temperatures • Chaperones are ATPases

What do the 4 types of bonds do for the tertiary structure? (Note: this is a particularly difficult question)

• Hydrogen bonds between the R-groups of the individual AA • Ionic bonds → bond between oppositely charged ions (anions - and cations +) (stronger than H-bond) • Disulfide bridges → S-S-bond between 2 thiol groups (S-containing groups) in AA-R-groups (strongbond) • Hydrophobic interactions → between hydrophobic (nonpolar) AA-side groups (can be strong)

Review: Golgi apparatus, where is it and what is it's function?

• In cytoplasm of eukaryotes • Place of modification of proteins after synthesis • Packages and stores proteins

alternative splicing (what is it and is is post translational or post transcriptional?)

• Specific signal sequences in the mRNA are recognized by an enzyme called "small nuclear ribonucleoprotein particle" (snRNP) and removed from the transcript • Post-transcriptional modification

Describe the steps of the folding cycle. (there are 5 steps)

• The folding cycle requires ATP • The chaperone is bound to ADP • ADP/Chaperone-complex has a high affinity for unfolded proteins • After binding ADP is released • After protein folding and ATP binding the complex dissociates and the correctly folded protein is released

POST-TRANSLATIONAL MODIFICATIONS OF PROTEINS, NAME THE FUNCTION

• This process describes modifications of many eukaryotic proteins to form mature proteins, helps to regulate enzyme activity, promotes folding, and aids with stabilizing proteins • It can extend the 20 standard AA by modifying their functional groups or by adding new ones, such as phosphate groups