Joachim w deitmer fb biologie mit fb mathematik und itwm
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Joachim W. Deitmer, FB Biologie mit FB Mathematik und ITWM. Die Rolle der Neuroglia bei der Bildung, Funktion und Plastizität von Synapsen. Räumlich-zeitliche Interaktionen zelluläre Signalmoleküle.

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Joachim W. Deitmer, FB Biologie mit FB Mathematik und ITWM

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Joachim w deitmer fb biologie mit fb mathematik und itwm

Joachim W. Deitmer, FB Biologiemit FB Mathematik und ITWM

Die Rolle der Neuroglia bei der

Bildung, Funktion und Plastizität

von Synapsen

Räumlich-zeitliche Interaktionen

zelluläre Signalmoleküle


Joachim w deitmer fb biologie mit fb mathematik und itwm

A model reduction approach to the kineticsof the monocarboxylate transporterMCT1 and carbonic anhydrase II

MCT

MCT

1 H+

1 Lac-

Joachim Almquist1, Henning Schmidt1, Patrick Lang2, Dieter Prätzel-Wolters2, Joachim W. Deitmer3, Mats Jirstrand1, and Holger Becker3

1Fraunhofer-Chalmers Centre, Gothenburg, Sweden

2Institut für Techno- und Wirtschaftsmathematik (ITWM) Fraunhofer Gesellschaft, Kaiserslautern, Germany

3Technische Universität Kaiserslautern, Kaiserslautern, Germany

Neuron-glia interactions


Joachim w deitmer fb biologie mit fb mathematik und itwm

Aim of this Project

To determine the mechanism of the monocarboxylate transporter (MCT1) and to present a mechanistic hypothesis of how MCT1 interacts with the enzyme carbonic anhydrase II (CAII). The modelling process might provide ideas for this.

To derive a rate expression for the MCT1 that also includes the effect of CAII. This could be used in other models.

Protein expression in Xenopus oocytes

Injection of rat MCT1-cRNA

Injection of CAII (isolated from bovine erythrocytes)

Microelectrodes for intrcellular pH measurements

ODE modelling

Model reduction

Methods - Electrophysiological Techniques and Mathematical Modeling


Functionally expressed proteins in xenopus oocytes interactions with carboanhydrases

Functionally expressed proteins in Xenopus oocytes: Interactions with carboanhydrases?

  • Messung von Membranströmen in ‚Voltage-Clamp‘

  • Messung von cytosolischem pH und Na+ mit ionen- selektiven Mikroelektroden

  • Struktur-Funktion-Analyse durch gezielte Mutationen

Funktionelle heterologe Expression

von Membrantransportproteinen in

Frosch-Oozyten (Xenopus laevis)

Carboanhydrase II

Modell der Wechselwirkung zwischen Carboanhydrase und

Membrantransporter (NBCe1=Natrium-Bikarbonat-Kotransporter

NBCe1 und Carboanhydrase ko-exprimiert


The model

The Model

Monocarboxylate

transporter

MCT

1 H+

1 Lac-

Ordinary differential equation model of the transporter states shown in the cartoon


Caii included in model

CAII Included in Model

The effect of CAII is included in the model as an increased rate of proton uptake and release on the intracellular side of the transporter.


Comparing measurements with simulations

Comparing Measurements with Simulations

Efflux experiments with and without CAII (A,B) are compared with the model (C).

Influx experiments with and without CAII (A,C) are compared with the model (B,D).


Kooperationen

Kooperationen

  • Prof. D. Prätzel-Wolters, FB Mathematik und ITWM

  • ITWM: Dr. P. Lang

  • Fraunhofer-Chalmers-Centre, Göteborg, Schweden: Prof. M. Jirstrand, J. Almquist

    Erfolge/Fortschritte: Erstes Paper über das Modell in Revision

    Bisher dem Projekt zugewiesene Mittel: 30 T€ für 2008


Weiterer fahrplan

Weiterer Fahrplan

  • Erweiterung des Modells mit Voraussagen (Einbeziehung von verschiedenen Isoformen und Mutanten der Carboanhydrase sowie mit NBCe1

  • Experimentelle Überprüfung dieser Voraussagen und Simulierung weiterer Parameter

  • Neues Projekt (DFG-Antrag wird gerade geschrieben): Mechanismen und Modellierung der Protonen-Pufferung in Zellen


Future projects

Future Projects

  • Measuring, analyzing and modelling of the capacity and dynamics of cellular H+ buffering

    - Spatial dynamics of buffering within a cell and role of carbonic anhydrases

    - The role of acid/base-coupled membrane transporters, such as the NBC, for buffering


Thank you

Thank you!

Gilt das nur für den Betze?


Measurement of buffer capacity

Measurement of buffer capacity

CA

CO2 +H2O

H+ + HCO3-

Henderson- Hasselbalch equation:pH = pK‘ log [HCO3-]/[CO2]

[HCO3-]i = 10(pHi-pK‘)x [CO2] (pK‘=6.1)

Buffer capacity = acid/base injection / dpHi

βt = intrinsic + CO2/HCO3—dependent

ßCO2 ≈ 2.3 [HCO3-]

dpHi

By measuring pHi, ß can be determined!

Addition/injection of acid


Model reduction

Model Reduction

One possible set of constraints that can be used to reduce the ODE-system. Solving this set of equations yields a explicit rate expression for the cross-membrane transport of MCT1 substrates,T.


Substrate inhibition predictions

Substrate Inhibition Predictions

Model reduction with different assumptions on transporter properties leads to predictions of inhibition by single substrate presence.


A hypothesis for the mct1 caii mechanism

A Hypothesis for the MCT1-CAII-mechanism

A hypothesis for the MCT1-CAII-mechanism. One or several CAII molecules close to the inner mouth of MCT1 might be working as a proton antenna. If proton uptake and release are the rate-limiting steps of transport, MCT1 turnover could be increase by this antenna.


Voltage dependence of the total buffer capacity t of oocytes expressing nbc

Voltage dependence of the total buffer capacity (ßT) of oocytes expressing NBC

Becker & Deitmer (2004) J. Biol. Chem.279, 28057-28062


Aim of these studies

Aim of these studies

  • Measuring and modelling (simulating) H+ buffer capacity

  • Predicting parameters of cellular buffering

  • Testing predicted parameters in experiment

  • What consequences do our findings might have for pH-dependent processes in systems (cells, tissue, organs)?


Data from substrate inhibition

Data from Substrate Inhibition

The inhibition predictions are tested in experiments.


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