Kiralj R., Ferreira M. M. C., “QSAR study of structurally unrelated substrates of MDR efflux pump VmrA from V. parahaemolyticus”. São Pedro, SP, 20-23/11/2005: XIII Simpósio Brasileiro de Química Teórica (XIII SBQT) [13th Brazilian Symposium of Theoretical Chemistry], Livro de Resumos [Book of Abstracts], 199. Poster 199.

199
QSAR study of structurally unrelated substrates of MDR efflux pump VmrA from V. parahaemolyticus

Rudolf Kiralj (PQ), Márcia M. C. Ferreira (PQ). rudolf@iqm.unicamp.br

Instituto de Química, Universidade Estadual de Campinas, 13083-970 Campinas, SP, Brazil

Palavras-Chaves: mutidrug resistance,  semi-empirical methods,  quantitative structure-activity relationships
(QSAR)

INTRODUCTION
Multidrug resistance  (MDR)  of microbes and parasites  as  well as of cancer cells to  currently used drugs  is
becoming one of the major problems in combating infectious and  parasitic diseases and cancer,  respectively.
Among major mechanisms  of  multidrug  resistance  in  cellular microbes  and cancer  cells  are  efflux pumps,
macromolecular systems that extrude drugs and  a large variety of structurally dissimilar substances  from cell
into the outside medium.   Bacterial  MDR efflux pump VmrA exists in  V. parahaemolyticus,  a marine bacteria
that causes frequently food poisoning in Japan and many other countries  [1].  VmrA  is being effective against
 several structurally unrelated drugs, organic dyes, detergents and xenobiotics.  It is a Na⁺/drug antiporter that
extrudes the substrates from  the  cytoplasm  to the periplasmic space  in  a  Gram-negative  bacterial cell.  Its
functional form consists of  a protomer placed in the inner membrane,  with  448  [1]  or  447  [2]  residues that
form twelve hydrophobic transmembrane segments  of  21-24  residues.  3D structure of VmrA is not know yet.
Substrates of this pump,  although  structurally  very  diverse,  must  share  some  common properties that are
responsible for their efflux from V. parahaemolyticus.
METHODS
In this work,  the   relationships   between  drug  molecular  properties  and  efflux  activity  of  VmrA  (negative
logarithm  of  Minimal Inhibitory  Concentration)  [1]   have  been  studied  by  means  of  quantitative structure-
activity relationships  (QSAR).  Twelve  drugs were included in the study:  DAPI  (4’,6-diamino-2-phenylindole),
TPPCl   (tetraphenylphosphonium   chloride),    acriflavine    chloride,    ethidium    bromide,   chloramphenicol,
norfloxacin,  rhodamine  6G  chloride,  tetracycline,   erythromycin,  streptomycin,  sodium  deoxycholate,  and
sodium  dodecyl sulfate.   Structures  of  the  active  organic  parts  of  all  drugs  were  modeled  according  to
available experimental structures for these  or  the most similar  drugs  in  the  Cambridge Structural Database,
taking into  account  the  ionic state  of  these  species  at neutral  pH.  The geometry  of  the neutral and ionic
species was optimized at  PM3  semi-empirical level in  Titan package.  Molecular descriptors were calculated
from    the  obtained   structures   by  using  Titan,   MOPAC 6.0   and   Chem3D   programs.  Other  molecular
descriptors, mainly of compositional  and  topological nature,  were  generated from two-dimensional chemical
formula.  The molecular descriptors data were autoscalled  prior  to  chemometric analysis that was performed
using  programs  Pirouette   and   Matlab.   Cut-off   0.50   in   correlation   coefficients   for   descriptor-activity
relationships was used in variable  selection.  PLS  (Partial  Least  Squares)  regression  and  PCR  (Principal
Component  Regression)  models  were  built  and validated by leave-one-out crossvalidation.
RESULTS AND DISCUSSION
The obtained  PLS and  PCR  regression  models  with  one  principal  component  (70% of the total variance)
resulted in very similar and satisfactory statistics: correlation coefficient of validation  Q > 0.80  and  prediction
R > 0.88,  and  standard error of  validation  SEV < 0.73  and  prediction  SEP < 0.57.  This  is  reasonable  to
expect since the twelve drugs belong to twelve different classes of compounds.  The first principal  component
discriminates the drugs according  to  their  biological  activities  with  respect  to  VmrA  pump.   DAPI,  TPPCl,
acriflavine  and  ethidium  bromide  behave  as  good  substrates  of  this  pump,  what  is  in  agreement  with
experimentally observed elevated resistance of V. parahaemolyticus to these drugs  [1]. The other drugs form
two groups that are partially mixed, as moderately good and poor substrates,  to  which there is a very modest
and  none  MDR  in  the  bacterial  cells,  respectively.  Good   substrates  are  characterized  by   rather  rigid
structures (planar fragments and rings)  and  predominant  hydrophobic character,  modest  polarizability  and
dipole  moment,  and   limited  content   of   polar   groups.   Certain   molecular  properties   exhibit   parabolic
relationships with the efflux activity of  VmrA,  thus indicating the optimal ranges of molecular descriptors   that
characterize  the  best and worst substrates of this  pump.  These  observations  agree  with  the  pronounced
hydrophobic character of this transmembrane pump.
CONCLUSIONS
Steric, eletronic and hydrophobicity molecular descriptors of twelve unrelated drugs are quantitatively related
to the efflux rate of the drugs, as extruded by pump VmrA in cells of V. parahaemolyticus.
ACKNOWLEDGEMENT: FAPESP
LITERATURE
[[1] J. Chen et al., J. Bacteriol. 184 (2002) 572-576.
[2] K. Makino et al., Lancet361 (2003) 743-749.