Introduction to Bioorganic Chemistry and Chemical Biology
Answers to Chapter 9 (in-text & asterisked problems)
Answer 9.1 Names and acronyms from August 2011 issues of chemistry/biology journals are listed below. The important point is that most of the papers in the current research literature involve mysterious acronyms. The goal of this chapter is to familiarize you with those acronyms that relate to signal transduction pathways that affect gene transcription in humans.
Journal of Biological Biological Chemistry 286 286,, Issue 34 (2011) Smad2/3 (a hybrid of C. elegans Sma (small) gene and Drosophila Mad (Mothers (Mothers against dpp) gene) Hepatocyte Nuclear Factor 4 α RNA Photosystem II (PSII)–Psb27 (photosystem protein b27) protein complex ATP synthase dentin matrix protein integrin MSH2 (DNA repair proteins homologous to yeast mutS /hexA) Cdt2 (protein target of target of Cdc10 transcriptional regulation) P21Waf1/Cip1 (kinase inhibitor protein 21 encoded by the Waf1 /Cip1 genes) VAMP8 VA MP8 (Vesicle-associated ( Vesicle-associated Membrane Protein 8) SNARE (Soluble N -Ethylmaleimide-sensitive -Ethylmaleimide-sensitive Factor Attachment Protein Receptor) GSK3β (Glycogen Synthase Kinase-3 β) PTTG1/Human Securin (protein from Pituitary Tumor-Transforming Gene 1 ) Myocyte Enhancer Factor 2C, an Osteoblast Transcription Factor aurora-A augmin Hice1 Protein (Hec1-interacting and centrosome-associated 1) microtubule ARD1 ( Arabidopsis Acireductone Dioxygenase 1) Heterotrimeric G protein β subunit Hsp90 (Heat-shock protein 90) Immunophilin FKBP51 (FK-506 binding protein 51) PSKs/TAOKs PSKs/T AOKs (Prostate-derived Sterile 20-like K inases) glucocorticoid Dig2/RTP801/REDD1 (dexamethasone-induced Gene 2 Protein) NKCC2 (bumetanide-sensitive Na +,K +,2Cl– Cotransporter) Tamm–Horsfall protein protein CaV1.3 channels (voltage-gated calcium channel 1.3) c-Jun kinase (the Jun oncogene from Avian sarcoma virus was named after Ju nana, the Japanese word for 17) Runx2 protein (Runt-related transcription factor 2) DinD protein (DNA damage inducible protein D) Escherichia coli DinD RecA (DNA recombinase A) VAP-1 VAP -1 ( Vascular Adhesion Protein-1) oxime acetylcholinesterase
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Introduction to Bioorganic Chemistry and Chemical Biology: ANSWERS TO CHAPTER 9
cytochrome P450 SPα (a heme enzyme that absorbs light at 450 nm) fatty acid autotaxin Egr-1 protein (Early Growth Response 1) HNF4α (Hepatocyte Nuclear Factor 4 α) SHP (Small Heterodimer Partner) Mef2A transcription factor (myocyte enhancer factor 2A) GRTH/DDX25 (Gonadotropin-regulated Testicular RNA Helicase) StAR (Steroidogenic Acute Regulatory protein) androgen receptor TGF-β1 (Transforming Growth Factor-β1) microRNA 143/145 TLR4 protein (Toll-like receptor 4) MMP-1 (Matrix Metalloproteinase-1)
Chemical Biology 50, Issue 8 (2011) retinal mitochondrial Lon protease docataxel tubulin Hsp90 (heat shock protein 90) JNK (Jun kinase) Sab kinase (SH3 domain-binding protein that preferentially associates with Btk) mycobacterial arabinosyltransferase decaprenylphosphorylarabinose analogues Kalata B1 K ir2 inward rectifier potassium channels (inwardly rectifying potassium channel 2) RNA
Biochemistry 50, Issue 33 (2011) p53 (mammalian tumor suppressor protein (53 kDa)) group 3 LEA protein (Late Embryogenesis Abundant) nucleoplasmin histone prion P. aeruginosa type III secretion translocators P. aeruginosa heteromeric transmembrane pores tyrosyl tRNA synthetase nitric oxide synthase Hsp70 (heat shock protein 70) thioredoxin rhodopsin phoborhodopsin prolyl–acyl carrier protein oxidase P. aeruginosa isochorismate–pyruvate lyase GTPase chloroplast signal recognition particle S100A4 (calcium-binding proteins soluble in 100% saturated ammon ium sulfate) myosin-IIA mouse olfactory receptor glmS riboswitch (glucosamine-6-phosphate activated ribozyme) glucosamine 6-phosphate α-anomer DNA polymerase RNA frataxin cyclotide GluA2 AMPA receptor (A2 ion channel receptor that binds to glutamate)
Introduction to Bioorganic Chemistry and Chemical Biology: ANSWERS TO CHAPTER 9
Science 333, August 11 (2011) Fcγ receptor (receptor that binds to immun oglobulin constant fragment γ) agonistic CD40 antibodies (complementarity determining/determinant 40)
Nature 476, Number 7360 (2011) Lgr5 homologues (leucine-rich repeat-containing G protein-coupled receptor 5) Wnt receptors (Wnt is a hybrid of the Drosophila Wingless gene and the mammalian integration 1 gene) R-spondin histone 14-3-3 proteins rice Hd3a florigen (heading date 3a) MCU (mitochondrial calcium uniporter) ribosome Answer 9.2 Cycloheximide would have no effect since mucin release does not involve protein synthesis. Physostigmine would enhance the effect of acetylcholine by preventing hydrolysis by acetylcholinesterase. Atropine would inhibit the rapid release of mucin by antagonizing the effect of acetylcholine. Answer 9.3 Nuclear receptors and gas receptors. Answer 9.4 The androgen receptor (AR) binds duplex DNA with the sequence AGGTCA; the estrogen receptor (ER) binds to duplex DNA with the sequence TGTTCT. By connecting the DNAbinding domain of the androgen receptor to FKBP12 and the DNA-binding domain of the estrogen receptor to mTOR, the two domains will dimerize and bind to DNA in the presence of rapamycin. Semiflexible peptide linkers would be needed to connect the domains; the C termini of FKBP12 and mTOR are closer together than the N termini. 5'-AGGTCANNNTGTTCT-3'
5'-AGGTCANNNTGTTCT-3'
3'-TCCAGTNNNACAAGA-5'
3'-TCCAGTNNNACAAGA-5'
AR DBD
ER DBD
rapamycin
AR DBD
ER DBD
FKBP12 mTOR
FKBP12
mTOR
Answer 9.5 The calculations suggest that roughly more than 1% of enone steroids exist as thiol adducts, but Nature is cautious. The equilibrium constant for addition to steroid enones such as testosterone is actually 100-fold less favorable ( K eq = 0.02) than it is for 3-methylcyclohexenone.
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Introduction to Bioorganic Chemistry and Chemical Biology: ANSWERS TO CHAPTER 9
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1. How much adduct, x , is present at equilibrium? K eq
HS
+ O
R'
R
[A]
If thiol is depleted:
10-6 M
O
[R'SH] - x
If thiol is not depleted: 10-6 M - x
S
R
thiol adduct
R'
[A•R'SH]
0.005 M - x
x
0.005 M
x
2. Set up equations depending on your assumptions
If thiol is depleted:
If thiol is not depleted: K eq
[A•R'SH]
=
[A] [R'SH]
=
( x ) (10 -6 M
- x)(0.005 M)
(10 -6 M - x )(0.005 M)
3. Plug in
1 K eq
1 K eq
x
)
O
progesterone adduct
- x )(0.005 M - x )
1 K eq
)• x + x 2 = 0
O
S
O
less reactive K eq ~
( x ) (10 -6 M
5x10-9 M2 - (0.005 M + 10-6 M +
O
x =
[A] [R'SH]
=
K eq
major prednisone adduct
R'
[A•R'SH]
... you end up with a quadratic equation; but since [R'SH] >> [A], the answers you calculate, will not be very different from the thiol depletion scenario
K eq
5x10-9 M2 (0.005 M +
=
x
=
5x10-9 M2 - (0.005 M)• x =
x =
K eq
S K eq ~
7,000 M-1
0.0097x10-6 M thiol adduct 0.990x10-6 M prednisone left over
x =
99% of progesterone left over
R' 2
0.0099x10-6 M thiol adduct 0.990x10-6 M progesterone left over 99% of progesterone left over
Answer 9.6 H435F, H435K, H435W, and H435R are too big and or too polar to form complementary interactions with the bulky ethoxy group of QH2. H435G is probably too small and would leave empty space. The H435A mutant is just right to complement the thyroid mimic QH2. Answer 9.7 A
IL-3, GM-CSF, IL-6, and IL-5
B
IL-3, IL-7, IL-6, and IL-5
Answer 9.8 Most noncovalent interactions between small molecules and proteins have fast on-rates and reach equilibrium very quickly. The fact that inhibition increases with longer incubation times and that the sterically similar analog is inactive suggests a covalent reaction between stattic and STAT3, which has numerous free cysteine residues. Cys712 (PDB 1BG1) makes contact with the active site and could form an adduct with the vinylsulfone moiety. Cys -
Cys
S
S
..
O2 N
S O O
O2N
S O O
