This PhD proposal outlines a study of the contribution of outer membrane permeability to antibiotic resistance in various bacterial organisms. Specifically, it will characterize the porin Omp35 from Enterobacter aerogenes and study its interaction with antibiotics and divalent ions like magnesium. Single channel recordings will examine Omp35 conductivity at different pH levels and its association rates with various antibiotics compared to its homolog OmpF. The effect of magnesium binding and how it modifies antibiotic binding kinetics will also be analyzed. Future plans include molecular dynamics simulations to study residue ionization in Omp35 and examine the docking sites of magnesium in other porins. This work seeks to further understand the specific uptake mechanisms of molecules by porins and their homologs.

1. Contribution of Outer Membrane Permeability of Various
Bacterial Organisms towards Antibiotic Resistance
Study of Porin Homologs and Divalent Ions Interaction
Committee Members:
Prof. Dr. Mathias Winterhalter
Prof. Dr. Ulrich Kleinekathöfer
Prof. Dr. Roland Benz
Dr. Mohamed Kreir
Jiajun Wang
PhD Proposal Defense

2. 2
7:50 PM
Material and Methods
Interaction
Kinetics
Residue Ionization
of Omp35
Introduction
Divalent ion
• OmpF, OmpC, Omp35 and Omp36 will be tested in this work.
• OmpF and OmpC are the outer membrane proteins found in
Escherichia coli and regulated according to osmotic pressure.
• Omp35 and Omp36 are the outer membrane proteins found in
Enterobacter aerogenes.
• OmpF and Omp35 are homologues and share 95% similarities in
the gene sequence.
• OmpC and Omp36 are homologues and share 90% similarities in
the gene sequence.
Future Plan
Questions Cowan, S. W., et al. (1992), Bornet C., (2004)

3. 3
7:50 PM
Characterization of Omp35
Interaction
Kinetics
Residue Ionization
of Omp35
Introduction
Divalent ion
• Single channel recordings of a single Omp35 porin incorporated
into an artificial lipid membrane at different pH value.
• Comparison between Omp35 and its homologue OmpF.
Future Plan
Questions

4. 4
7:50 PM
Molecule Interaction with Omp35
Interaction
Kinetics
Residue Ionization
of Omp35
Introduction
Divalent ion
• Current recordings of interaction between Omp35 and imipenem,
ertapenem, cefepime, ceftazidime and enrofloxacin.
• Association rate comparison between Omp35 and its homologue
OmpF.
Future Plan
Questions

5. 7:50 PM
Analysis on the Effect from Magnesium
5
Interaction
Kinetics
Residue Ionization
of Omp35
Introduction
Divalent ion
• Binding region of Magnesium ions.
• Enrofloxacin, Norfloxacin binding kinetics modified by Mg2+.
• Mg2+ modify kinetics between norfloxacin and other porin
homologs.
Future Plan
Questions

6. 6
7:50 PM
Future Plan
Interaction
Kinetics
Residue Ionization
of Omp35
Introduction
Divalent ion
Future Plan
Questions
OmpF and Omp35
- Ertapenem, Imipenem,
Cefepime, Ceftazidime,
Enrofloxacin, Norfloxacin.
Residue Ionization
- Conductance and I-V response of
Omp35.
- Conductance vs. pH value
Molecular Dynamic Simulation
- Check if the computer simulation
give the same residue ionization
in Omp35, Omp36 etc.
Porin Homologs and Mutagenesis
- These molecules will be checked
with OmpC and Omp36. Omp37,
OmpK35, OmpK37.
- Liposome swelling assay.
Magnesium Ions
- Strong effect on the
fluoroquinolone in OmpF.
- Slightly changed the kinetics
with carbapenem and
cephalosporin.
MD Simulation and Mutagenesis
- Docking site of Mg2+ in other
porins.
- Mg2+ effect the interaction
mechanism.
Cation Ions
- Mg2+ has been checked.
Ions
- Ca2+, SO4
2-, F- etc. can also be
checked
Specific Channels Uptake Mechanism General Porins of Specific Molecule Uptake

7. 7
7:50 PM
Acknowledgement
Interaction
Kinetics
Residue Ionization
of Omp35
Introduction
Divalent ion
Future Plan
Questions
Prof. Dr. Mathias Winterhalter
Dr. Niels Fertig
Dr. Mohamed Kreir
Harsha Bajaj
Satya Prathyusha Bhamidimarri
All the members of Winterhalter group
Nanion Technologies GmbH