532 - Med Chem Ii - Exam I - Part 1, 2A, & 2B Questions

Front 1

List three advantages of solid phase peptide synthesis over the classical solution-phase method.

Back 1

1) The solid phase peptide synthesis allows reactions to be driven to completion by using a huge excess of reagents, 2) The rinsing of the solid phase resin after each chemical step makes purification easy, & 3) There are a widely diverse selection of resins

Front 2

List three disadvantages of solid phase peptide synthesis over the classical solution-phase method.

Back 2

1) It utilizes vast amounts of solvent, compared to the classic method, 2) It is more limited in scale than an "all solution" process, & 3) The fidelity of solid phase peptide synthesis becomes suspect with peptides > 50 amino acids long

Front 3

What is the process by which one peptide, having a synthetic segment containing a thioester on an alpha-carbonyl group, undergoes nucleophilic attack by the cysteine residue at the amino terminal of another peptide, initially leading to a thioester ligation product that undergoes intramolecular reaction to yield a peptide bond at the ligation site?

Back 3

Native Chemical Ligation (NCL), a.k.a. Kent Ligation

Front 4

How can the Native Chemical Ligation process be modified to yield a peptide lacking the cysteine side chain?

Back 4

The Native Chemical Ligation sequence is followed by a desulfurization step, using hydrogen gas and palladium catalyst, that converts the cysteine side chain to a simple alanine residue by replacing the thiol group with a hydrogen.

Front 5

What are the advantages of Native Chemical Ligation over Standard Peptide Synthesis?

Back 5

1) Large peptides (35-60 aa) can be effectively coupled, 2) Coupling does not require sidechain protection, & 3) Coupling can be done in water at pH ~7.0

Front 6

What are the advantages of Native Chemical Ligation (NCL) over Biological Protein Production

Back 6

1) NCL is free of inherent biological contaminants (nucleic acids, viruses, prions, etc.), 2) NCL is easier to "scale up" than biological expression methods, 3) NCL can easily introduce "unnatural (uncoded)" amino acids into proteins, 4) NCL allows specific labeling of protein at unique sites with natural or unnatural groups, 5) NCL can be used to make circular proteins having enhanced biostability in vivo, 6) D-amino acids can be used to make proteins of the "opposite" stereochemistry from what is seen in nature. Such proteins could have vastly improved half-lives in vivo.

Front 7

Provide two reasons why the fidelity of solid phase peptide synthesis becomes suspect beyond ~50 amino acids.

Back 7

1) In spite of the 99% yield for each step, the overall yield is ~50% upon reaching a length of 50 amino acids & 2) As the peptide lengthens, the formation of secondary and tertiary structures may lower the yield of subsequent amino acid additions.

Front 8

How do factors such as temperature, pH, salt concentration, etc. influence the stability of peptide drugs?

Back 8

Factors such as the temperature, pH, and salt concentration influence the physical stability of secondary, or even higher order, structures essential to a peptide drug's activity.

Front 9

What are four processes associated with chemical instability in the primary structure of peptide drugs?

Back 9

1) Racemization, 2) Hydrolysis, 3) Beta-Elimination, & 4) Oxidation

Front 10

Which two amino acid side chains are susceptible to hydrolysis, and what side chains to they become upon hydrolysis?

Back 10

The amide functional groups on the side chain of the amino acids asparagine (asn) and glutamine (gln) are susceptible to hydrolysis, becoming aspartic acid (asp) and glutamic acid (glu), respectively.

Front 11

Which three amino acid side chains have functional groups making them susceptible to beta-elimination reactions?

Back 11

Cystine (cys) has a thio group suitable for beta-elimination, while serine (ser) and threonine (thr) both have a hydroxy group suitable for beta-elimination.

Front 12

Which two amino acid side chains have functional groups susceptible to oxidation?

Back 12

Cystine (cys) has a thio group (-SH) that may be oxidized, while Methionine (met) has a sulfide group (-SCH3) that may be oxidized.

Front 13

Name for a group of enzymes that catalyze the breakdown (hydrolysis) of the peptide bonds of proteins (or peptides).

Back 13

Proteases (or Peptidases)

Front 14

How do proteases capable of limited proteolysis differ from those capable of unlimited proteolysis?

Back 14

Proteases capable of limited proteolysis can only break specific peptide bonds, while those capable of unlimited proteolysis can completely break down a peptide.

Front 15

Explain the difference between an endopeptidase and an exopeptidase.

Back 15

Endopeptidases cleave the middle of a peptide, while exopeptidases cleave the terminal amino acid of the substrate.

Front 16

There are two types of exopeptidases. What are they and how does their active site dictate from which end of the peptide they cleave an amino acid.

Back 16

Aminopeptidases generally have residues with negatively charged side chains in the active site to bind the positively charged amino terminus, while carboxypeptidases generally have residues with positively charged side chains to bind the negatively charged carboxyl terminus.

Front 17

What do serine, cysteine, and threonine proteases have in common?

Back 17

Serine, cysteine, and threonine proteases are all covalent catalysts that catalyze amide hydrolysis indirectly.

Front 18

What do aspartic acid and metalloproteases have in common?

Back 18

Aspartic acid and metalloproteases are both general acid-base catalysts that catalyze amide hydrolysis directly by activating a water molecule.

Front 19

Describe the convention for numbering the substrate side chains and protease subsites.

Back 19

Beginning from the sites on each side of the scissile bond, the non-primed subsites are numbered upwards towards the N terminus (S1, S2, S3, etc for the substrate & P1, P2, P3, etc. for the protease), while the primed subsites are numbered upwards towards the C terminus (S1', S2', S3', etc. & P1', P2', P3', etc for the substrate and protease, respectively).

Front 20

What is a zymogen?

Back 20

A zymogen is an inactive enzyme precursor (a pro-enzyme). It requires a biochemical change, either a hydrolysis reaction or a conformational change, to reveal the active site.

Front 21

Describe the mechanism of a protease that functions via covalent catalysis. [List three specific examples of proteases of this class].

Back 21

Covalent catalysis involves indirect amide hydrolysis. A nucleophilic amino acid side chain of the protease forms an acyl-enzyme intermediate with the peptide substrate, which hydrolyzes to cleave the amide and free up the protease. [Serine Proteases, Threonine Proteases, & Cysteine Proteases]

Front 22

Describe the mechanism of a protease that functions via general acid-base catalysis. [List three specific examples of proteases of this class].

Back 22

General acid-base catalysis involves direct amide hydrolysis. The water molecule is activated by binding in a small cleft defined by two aspartic acid (or glutamic acid) residues of a protease, or by ligation to zinc. [Aspartic Acid (or Glutamic Acid) Protease is the name for the former, while Zn-metalloproteases are the name for the latter].

Front 23

What does the acronym serpins stand for? Why are they important? And what makes them unique among the many families of serine protease inhibitors?

Back 23

The acronymic name SERPINs stands for SERine Protease INhibitor proteins. Serine proteases are unique in that they rely highly on these proteins for regulation. Serpins are unique in that they possess a mobile reactive site loop having the ability to profoundly change its conformation to facilitate the formation of a virtually irreversible complex with their target proteases.

Front 24

Why does the serine protease inhibitor phenylmethylsulfonylfluoride (PMSF) only react with the serine residue in the active site of serine protease and not with other serines?

Back 24

PMSF only reacts with the serine residue in the active site of serine protease, and not with other serines, because the one in the active site is part of a catalytic triad of residues that serve to activate the serine oxygen, increasing its nucleophilicity.

Front 25

What is the difference between a reversible and an irreversible cysteine protease inhibitor?

Back 25

A reversible cysteine protease inhibitor, containing an aldehyde or nitrile for instance, forms a covalent but reversible intermediate with the cysteine residue, while an irreversible cysteine protease inhibitor permanently modifies the protease via an irreversible alkylation or acylation of the active site.

Front 26

What serine protease cleaves peptide amide bonds where the carboxyl side of the amino acid (P1) is a lysine or arginine, except when either is followed by proline?

Back 26

Trypsin

Front 27

What serine protease cleaves peptide amide bonds where the carboxyl side of the amino acid (P1) is a tyrosine, tryptophan, or a phenylalanine?

Back 27

Chymotrypsin

Front 28

What serine protease cleaves peptide amide bonds having a small, aliphatic P1 residue, such as alanine or valine?

Back 28

Elastase

Front 29

List three residue sequences that can function as a catalytic triad for a serine protease.

Back 29

1) Ser-His-Asp, 2) Ser-His-Glu, & 3) Ser-His-His

Front 30

What are the only enzymes that utilize a threonine residue as a catalytic nucleophile for amide bond hydrolysis?

Back 30

Proteasomes

Front 31

What aspartic protease released by the stomach functions to degrade food proteins into peptides?

Back 31

Pepsin

Front 32

What aspartic protease plays an integral role in mediating extracellular volume, arterial vasoconstriction, and, consequently, mean arterial blood pressure?

Back 32

Renin, a.k.a. angiotensinogenase, participates in the Renin-Angiotensin System that mediates extracellular volume, arterial vasoconstriction, and thus mean arterial blood pressure.

Front 33

What aspartic proteases play a vital role in mammalian cellular turnover processes, such as bone resorption?

Back 33

Cathepsins

Front 34

In what protease class is BACE (beta-amyloid converting enzyme)?

Back 34

BACE is an Aspartic Protease

Front 35

In what protease class is the HIV-1 protease?

Back 35

HIV-1 protease is an Aspartic Protease

Front 36

What is HAART and how did it impact HIV therapy?

Back 36

The acronym HAART stands for Highly Active AntiRetroviral Therapy. The usual HAART regimen combines three or more antiretroviral drugs and has reduced the amount of active virus to undetectable levels, in some cases. HAART dramatically reduced the number of deaths due to HIV.

Front 37

How does a viral reverse transcriptase inhibitor work?

Back 37

A reverse transcriptase inhibitors blocks the enzymatic function of reverse transcriptase, preventing it from completing the transcription of viral single stranded RNA into a double-stranded viral DNA.

Front 38

What is the active form of didanosine?

Back 38

2',3'-dideoxyadenosine triphosphate is the active form of didanosine.

Front 39

How are adefovir dipivoxil and tenofovir disoproxil structurally different from earlier NRTIs and why are they prodrugs?

Back 39

Adefovir dipivoxil and tenofovir disoproxil differ structurally from earlier NRTIs in that they have an open chain in place of the 5-membered ring. They are formulated as prodrugs to mask the phosphate group of their respective active metabolites in order to increase cell permeability for enhanced delivery.

Front 40

What are the main differences between NRTIs and NNRTIs in terms of structure and binding site?

Back 40

Nucleoside Reverse Transcriptase Inhibitors (NRTIs) structurally mimic natural nucleosides. Reverse transcriptase incorporates the NRTIs into the proviral DNA chain, where they cause premature termination of the sequence. Non-Nucleoside Reverse Transcriptase Inhibitors (NNRTIs) bind to an allosteric site adjacent to the substrate binding site of reverse transcriptase, causing a conformational change that inhibits the transcription process.

Front 41

What is the key structure feature of HIV protease?

Back 41

The key structural feature of HIV protease is that it is comprised of two identical protein units, giving it C2 symmetry.

Front 42

What are the P1 and P1’ residues of the substrate of HIV protease.

Back 42

The P1 residue of the substrate of HIV protease is either phenylalanine (phe) or tyrosine (tyr), while the P1' residue is proline (pro).

Front 43

What are TSIs and what is the rationale behind their design?

Back 43

TSIs are Transition State Inhibitors. TSIs mimic the transition state of the tetrahedral intermediate of the amide bond hydrolysis reaction catalyzed by the protease. By mimicking that transition state, the TSIs bind the protease active site, thus precluding it from interacting with the tetrahedral intermediate of the targeted substrate.

Front 44

What dipeptide was the starting point in the development of saquinavir and why?

Back 44

L-Phe-L-Pro - connected through a hydroxyethylamine transition state mimic - is the dipeptide starting point in the development of saquinavir because these are the P1 and P1' residues, respectively, of the HIV protease substrate.

Front 45

What is Kaletra and what is the rationale behind its development?

Back 45

Kaletra is a HIV protease combination that couples lopinavir with ritonavir. Though both drugs have HIV protease activity, the rationale behind the combination is that ritonavir inhibits CYP3A4, hindering its metabolism of lopinavir.

Front 46

How was the hybridization strategy used for designing Indinavir?

Back 46

The C2 symmetry of HIV-protease allowed the hybridization of the P' half of Saquinavir with the P' half of another lead compound to create the structure of Indinavir. The C2 symmetry of HIV-protease essentially allows P and P' halves of any two protease inhibitors to be coupled, in any combination, to create structures that will predictably have protease inhibitor activity.

Front 47

How is Tipranavir different from earlier PIs? What lead compound was used for Tipranavir development? What is the key interaction between Tipranavir and the active site of HIV protease?

Back 47

Tipranavir is different from earlier protease inhibitors in that it is the first not made of peptides, thus it lacks amide bonds. Warfarin was the lead compound in the development of Tipranavir. The key interaction is between the two aspartic acid residues of HIV protease with the hydroxy group of Tipranavir.

Front 48

What is the mechanism of action of Maraviroc?

Back 48

Maraviroc inhibits the cellular entry of HIV-1 by blocking the interaction between HIV-1 and the CCR5 co-receptor on host cells.

Front 49

How does Enfuvirtide (Fuzeon) inhibit HIV fusion?

Back 49

HIV fusion occurs when the loosely-structured HR2 domain of the viral gp41 protein coils onto the exposed grooves of the trimeric HR1 domain, pulling the viral and host cell membranes into close proximity. Enfuvirtide (Fuzeon) is homologous to part of the HR2 region. It binds to gp41, blocking the HR-2 zipping process, thus precluding fusion of the viral and host cell membranes.

Front 50

What is the target and mechanism of Raltegravir?

Back 50

Raltegravir targets HIV integrase, inhibiting a key step referred to as the "strand transfer step" of integration.