Structural and Biochemical Studies of LysM Proteins

Research output: Book/anthology/dissertation/reportPh.D. thesis

The Lysin Motif (LysM) is a well characterised carbohydrate-binding module that is present, usually as repeated entities, in proteins from all organisms except archaea. The functional significance of the multiplicity of the LysM module was investigated using two homologous NlpC/P60 endopeptidases involved in peptidoglycan hydrolysis; the Cell Wall Lytic enzyme associated with cell Separation (CwlS) from Bacillus subtilis, and P60_Tth from Thermus thermopiles. Biochemical studies conducted on purified CwlS showed that multiple LysM modules function cooperatively to bind N-acetylglucosamine (NAG) polymers, and that the increased affinity was positively correlated to the catalytic activity of the enzyme. The cooperativity of LysM domains was further demonstrated in structural studies on P60_2LysM, a variant of P60_Tth that contains only two LysM domains and lacks the catalytic domain. Ligand-induced intermolecular dimerization was observed in the co-crystal structure of P60_2LysM and NAG6. Until today, this is the only structural evidence illustrating intermolecular dimerization of LysM proteins.
Intermolecular dimerization of plant LysM receptor kinases (RK) has been proposed as a mechanism for the initiation of downstream signalling. Indeed, several LysM-RK pairs have been reported to be implicated in symbiosis and defence signalling. One partner of the LysM-RK pair usually lacks an active kinase domain and most probably recruits the other active LysM-RK as a co-receptor to mediate downstream signalling. An example of such a LysM-RK pair is the Nod factor receptors 1 and 5 (NFR1 and NFR5) from Lotus japonicus. Upon recognition of rhizobial Nod factors, NFR1 and NFR5 initiate symbiosis signalling, subsequently leading to the development of root nodules and symbiotic nitrogen fixation. Most of the signalling components in the Nod factor signalling pathway have been identified through genetic approaches. The current symbiosis signalling model, however, lacks components that could link Nod factor perception at the plasma membrane to downstream responses, such as calcium influx and perinuclear calcium spiking events that are required for the formation of infection threads and nodules. Using a novel proteomics approach, nine receptor kinases and four receptor-like cytoplasmic kinases were found in pull-down experiments using an NFR5 bait. These putative interactors are currently being validated as components of the NFR5 signalling complex and their functional roles are being characterised.
Original languageEnglish
Number of pages259
Publication statusPublished - 9 Jan 2017

Note re. dissertation

PhD defence 9 January 2017<br/>Supervisor: Søren Thirup

    Research areas

  • LysM, Intermolecular Dimerization, NFR5

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