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Import and export and secondary metabolite production

The rishirilid biosynthetic gene (figure) cluster from Streptomyces bottropensis Gö C4/4 was recently cloned, sequenced and successfully expressed in Streptomyces albus (1). The cluster contains four transporter genes. rslT1, rslT2 and rslT3 encode an ABC transporter system, rslT4 encodes a MFS multidrug transporter. A careful analysis revealed that RslT1 is similar to ABC transporter amino acid binding proteins containing a lipobox at residue 35 and 36 and a characteristic signal peptide, RslT2 is an ATP binding protein and RslT3 is a permease. Based on the putative function of RslT1 the ABC transporter system more likely imports an amino acid into the cell than exports rishirilid into the periplasmic space (2).

Figure: Structure of rishirilid A and organization of the rishirilid biosynthetic gene cluster

Although structures of several ABC transporters have been reported (cited in 3) not much is known about substrate binding domains of these transporters and about the mechanisms how ABC transporters select and translocate their substrates. Aside the ABC transporter system the rishirilid gene cluster contains rslT4 which most probably is involved in the export of rishirilid into the extracellular space. RslT4 belongs to the EmrB/QacA transporter family which is known to be drug:H+ antiporter with 12 transmembrane domains sharing conserved sequence motifs.

The main focus of the proposed project is to understand the exact function of RslT1, RslT2, RslT3 and RslT4 by structural and functional studies.

The specific aims of the proposal are:

  • perform studies to show that L-valine is involved in rishirilid biosynthesis
  • determine the substrate specificity of RslT1
  • determine the structure of RslT1 at high resolution with bound substrate (in cooperation with O. Einsle)
  • perform kinetic studies to study L-valine binding (in cooperation with S. Andrade)
  • determine the structure of the RslT2-RslT3 complex at high resolution with bound substrate (in cooperation with O. Einsle)
  • characterize the substrate specificity of RslT4



[1] X. Yan, K. Probst, A. Linnenbrink, M. Arnold, T. Paululat, A. Zeeck, A. Bechthold:  Cloning and heterologous expression of three type II PKS gene clusters from Streptomyces bottropensis (2012) Chembiochem 13(2), 224–30.
[2] E. Biemans-Oldehinkel, M.K. Doeven, B. Poolman: ABC transporter architecture and regulatory roles of accessory domains (2006) FEBS Letters 580(4), 1023–1035.
[3] J. Yu, J. Ge, J. Heuveling, E. Schneider, M. Yang: Structural basis for substrate specificity of an amino acid ABC transporter (2015) PNAS 112(16), 5243-5248.


Prof. Dr. Andreas Bechthold

Institute of Pharmaceutical Sciences
79104 Freiburg

Phone: +49 (761) 203 8370
Fax: +49 (761) 203 8383