Speziesunterschiede im organischen Kationentransporter OCT1: Vergleich der Effekte der Aminosäuren F159, W217 und D474 in OCT1 des Menschen, der Maus und der Ratte
Species-specific differences of the organic cation transporter OCT1: effects of the amino acids F159, W217 and D474 in OCT1 of the human, mouse and rat
von Maximilian Bolesta
Datum der mündl. Prüfung:2020-12-09
Erschienen:2020-11-26
Betreuer:Prof. Dr. Mladen Tzvetkov
Gutachter:Prof. Dr. Mladen Tzvetkov
Gutachter:Prof. Dr. Hubertus Jarry
Gutachter:Prof. Dr. Margarete Schön
Gutachter:Prof. Dr. Margarete Schön
Dateien
Name:Dissertation_Bolesta_final_eDiss.pdf
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Zusammenfassung
Englisch
OCT1 is an integral membrane transporter that is able to transport certain substrates with cationic components in the molecular structure across the plasma membrane of cells. This transporter is important for the human organism in the plasma membrane of the hepatocytes because here OCT1 is involved in the elimination of exogenous and endogenous substrates. OCT1 is polyspecific. Substrates with different molecular structure can be transported. Nevertheless the smallest changes in the molecular structure of the substrate causes that it is no longer recognized by OCT1 and is therefore no longer transported. The function of OCT1 as a polyspecific transporter is the subject of current research. Important goals are knowledge of relevant amino acids for substrate binding and translocation by the transporter. These findings let us understand better the polyspecificity of the transporter. For a long time Oct1 in rats (rOct1) served as a model for understanding OCT1 in humans (hOCT1) and mice (mOct1). In this work, the transport behavior of the three species was compared after mutating analogous amino acids in mOct1 and hOCT1, which have been identified as key amino acids for membrane transport for rOct1. The aim of this work was to compare the effects of the mutation of important amino acids and thus to conclude to what extent the key amino acids of rOct1 also play an important role in hOCT1 and mOct1. It was shown that analogous mutations of important amino acids in hOCT1 and mOct1 produced different effects. These effects in turn differed from the results on rOct1. The amino acid D475 of rOct1 is also important in hOCT1 and mOct1 for substrate transport and of great importance for its kinetics. However, the transport by hOCT1 is completely dependent on D474 whereas the transport of some substrates by mOct1 can happen without the negative charge of D475. The mutation D475N did not lead to a loss of transport but to a change to the high capacity-low affinity transport mode for TEA+ and MPP+. This shows that mOct1 transports substrates differently than hOCT1. Phenylalanine on codon 159 (human) or 160 (mouse) is more important in humans than in mice. The removal of the benzole residue after a mutation from phenylalanine to alanine led to more drastic changes in hOCT1 than in mOct1. The important role of W217 / 218 cannot be excluded with certainty because the results showed no significant difference to the wild type. In addition, the transport behavior of rOct1, mOct1 and hOCT1 without any mutation for the substrate ranitidine was analyzed. In this work it could be shown that the transport behavior differs significantly between rOct1, mOct1 and hOCT1. It is particularly interesting that the ranitidine transport showed functional similarities between rOct1 and hOCT1 although these two transporters differ greatly from one another in the amino acid sequence. The transport of these two species differed from mOct1 although the structure of rOct1 and mOct1 is almost the same. Taken together, the results of this work confirm that rOct1, mOct1 and hOCT1 show differences in the transport mechanism, which is reflected in the different transport strengths of different substrates. A direct transfer of the results from the mouse / rat animal model to humans is therefore not allowed. Ultimately, this prohibits the clinician from transferring results from animal models on the kinetics of OCT1 substrates to the collective of patients to be treated.
Keywords: cation; transporter; OCT1; hOCT1; mOct1; rOct1; Ranitidine; Sumatriptan; fenoterol; TEA+; MPP+; organic