Matthew  KimA,  Kristen  Procko*  and  Stephen  F.  Martin‡  

A  =  Synthesis  and  Biological  Recognition,  *  =  Research  Educator,  ‡  =  Principal  Investigator          

   The  main  goal  of  this  study  is  to  determine  the  thermodynamics  of  protein-­‐

ligand  interactions  to  the  stream’s  target  protein,  the  mouse  major  urinary  protein-­‐I  (MUP-­‐I),  which  functions  in  the  protection,  transport,  and  slow  release  of  pheromones.  Studying  MUP-­‐I  can  introduce  a  more  clear  understanding  of  the  unorthodox  non-­‐classical  hydrophobic  effect,  in  which  the  protein-­‐ligand  interaction  is  driven  enthalpically  as  opposed  to  entropically.  This  sparked  interest  in  structurally  changing  various  ligands  in  order  to  study  the  thermodynamic  behavior  arising  from  their  interactions  to  MUP-­‐I.  An  analogue  of  a  high-­‐binding  ligand,  2-­‐sec-­‐butyldihydrothiazole  (SBT),  was  synthesized.  It  was  predicted  that  because  of  the  analog’s  unique  structure,  an  increase  in  the  enthalpic  stabilization  could  occur,  leading  to  a  higher  binding  afRinity  than  that  of  SBT.    

General  characteristics:  §  Functions  in  the  protection,  transport,  and  slow  release  of  pheromones  §  Single  α-­‐helix  and  eight  β-­‐sheets  §  Forms  hydrophobic  barrel  §  Active  sites  hydrated  with  two  water  molecules  §  Exhibits  hydrophobic  effect  

Important  active  site  characteristics:  §  Hydrogen  bond  network  consists  of  Phe56,  Leu58,  and  Tyr138  §  Van  der  Waals  interactions  with  multiple  residues  in  the  active  site:  

§  Leu72,  Val100,  Phe108,  Leu121,  leu123,  and  Leu134    

 

•  Ligand  is  normally  introduced  incrementally  into  a  solution  of  MUP-­‐I;  the  heat  released  upon  binding  is  recorded.    

•  Free  energy  is  determined  through  values  derived  from  the  Gibbs  free  energy  equation,  ΔG  =  ΔH  –  TΔS  =  -­‐RT(lnKa).    

•  Change  in  free  energy  (ΔG),  which  is  inRluenced  by  the  change  in  enthalpy  (ΔH)  and  change  in  entropy  (ΔS),  reveals  the  driving  forces  for  a  particular  ligand’s  binding  to  the  active  site  of  MUP-­‐I.  

S

N

Leu72

Leu134

Phe108

Val100

Ala121

HO

HO H O

H

O

H O

Leu58

Tyr138

Leu123

NH

NH

Phe56

Abstract1  

Mouse  Major  Urinary  Protein  –  I1,3,4  

Observed  Trends4   Isothermal  Titration  Calorimetry  (ITC)2,3  

ITC  Data  

Acknowledgements  

References:  1Timm,  D.E.;  Baker,  L.J.;  Mueller,  H.;  Zidek,  L.;  Novotny,  M.V.  Structural  basis  of  pheromone  binding  to  mouse  major  urinary  protein  (MUP-­‐I).  Protein  Sci.  2001,  10,  997-­‐1004.  2Pierce,  M.M.;  Raman,  C.S.;  Nall,  B.T.  Isothermal  Titration  Calorimetry  of  Protein-­‐Protein  Interactions.  Methods  1999,  19,  213-­‐221.  3Homans,  W.S.  Water,  water  everywhere  –  except  where  it  matters?  Drug  discovery  today  2007,  12,  13-­‐14.  4Sharrow,  S.D.;  Novotny,  M.V.;  Stone,  M.  J.  Thermodynamic  analysis  of  binding  between  mouse  major  urinary  protein-­‐I  and  the  pheromone  2-­‐sec-­‐  butyl-­‐4,5-­‐dihydrothiazole.  Biochemistry  2003,  42,  6302-­‐6309.  5Jin,  Sarah.  The  University  of  Texas,  Austin,  TX.  4,5-­‐Dihydro-­‐2-­‐phenylthiazole,  2010.  6Azarm,  Kristopher.  The  University  of  Texas,  Austin,  TX.  Elucidating  the  Thermodynamics  of  Binding  of  2-­‐(Pyridin-­‐3-­‐yl)-­‐4,5-­‐dihydrothiazole,  2011.    

   N S HS

H2N

DBHReflux, 7 min. 100 oC

N S

N

4 5 6

Synthetic  Route  

S

N

S

N

1 2

S

N N

3

Past  Studies5,6