Vitamin B12 Transport in Bacteria: A structural and biochemical study to identify new transport systems

Stephan Rempel

Research output: ThesisThesis fully internal (DIV)

415 Downloads (Pure)

Abstract

Uptake of vitamin B12 is essential for many bacteria, but in most cases the membrane proteins involved in transport still need to be identified. Here, the biochemical characterization and high-resolution crystal structure determination of ECF-CbrT and BtuM, two predicted bacterial vitamin B12 uptake systems. ECF-CbrT belongs to the energy coupling factor (ECF-) type ABC-transporter superfamily, which use energy from ATP hydrolysis to power transport. ECF-transporter consist of four components, two that use ATP to fuel transport and one that acts, among other functions, as a scaffold, which together form the tripartite ECF-module. The fourth component acts as a membrane embedded substrate-binding protein. ECF-CbrT exhibits all typical features of an ECF-transporter and binds vitamin B12 with high affinity. Additionally, by measuring transport of vitamin B12 in a defined, reconstituted system allowed for the characterization of the overall transport reaction. The second protein presented here is BtuM, which has no similarity on the sequence level with any other known proteins, but surprisingly is structurally related to the substrate binding component of ECF-type transporters. In contrast to ECF transporters, BtuM does not require the other three components for transport. Therefore, BtuM represents a novel class of ECF-module independent transporters. Additionally, BtuM uses an unprecedented thiolate coordination to bind vitamin B12 that allows for chemical modification. The latter feature is extraordinarily rare among membrane transporters. The results presented in this thesis shed light on the diversity of transporters for vitamin B12 and pave the way for future research and anti-microbial drug design.
Original languageEnglish
QualificationDoctor of Philosophy
Awarding Institution
  • University of Groningen
Supervisors/Advisors
  • Slotboom, Dirk, Supervisor
  • Poolman, Berend, Supervisor
Award date18-Jan-2019
Place of Publication[Groningen]
Publisher
Print ISBNs978-94-034-1284-9
Electronic ISBNs978-94-034-1283-2
Publication statusPublished - 2019

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