TY - JOUR
T1 - Structural Analysis of the Hanks-Type Protein Kinase YabT From Bacillus subtilis Provides New Insights in its DNA-Dependent Activation
AU - Shi, Lei
AU - Cavagnino, Andrea
AU - Rabefiraisana, Jean-Luc
AU - Lazar, Noureddine
AU - Li de la Sierra-Gallay, Inés
AU - Ochsenbein, Francoise
AU - Valerio-Lepiniec, Marie
AU - Urvoas, Agathe
AU - Minard, Philippe
AU - Mijakovic, Ivan
AU - Nessler, Sylvie
PY - 2019
Y1 - 2019
N2 - YabT is a serine/threonine kinase of the Hanks family from Bacillus subtilis, which lacks the canonical extracellular signal receptor domain but is anchored to the membrane through a C-terminal transmembrane helix. A previous study demonstrated that a basic juxtamembrane region corresponds to a DNA-binding motif essential for the activation of YabT trans-autophosphorylation. YabT is expressed during spore development and localizes to the asymmetric septum where it specifically phosphorylates essential proteins involved in genome maintenance, such as RecA, SsbA, and YabA. YabT has also been shown to phosphorylate proteins involved in protein synthesis, such as AbrB and Ef-Tu, suggesting a possible regulatory role in the progressive metabolic quiescence of the forespore. Finally, cross phosphorylations with other protein kinases implicate YabT in the regulation of numerous other cellular processes. Using an artificial protein scaffold as crystallization helper, we determined the first crystal structure of this DNA-dependent bacterial protein kinase. This allowed us to trap the active conformation of the kinase domain of YabT. Using NMR, we showed that the basic juxtamembrane region of YabT is disordered in the absence of DNA in solution, just like it is in the crystal, and that it is stabilized upon DNA binding. In comparison with its closest structural homolog, the mycobacterial kinase PknB allowed us to discuss the dimerization mode of YabT. Together with phosphorylation assays and DNA-binding experiments, this structural analysis helped us to gain new insights into the regulatory activation mechanism of YabT.
AB - YabT is a serine/threonine kinase of the Hanks family from Bacillus subtilis, which lacks the canonical extracellular signal receptor domain but is anchored to the membrane through a C-terminal transmembrane helix. A previous study demonstrated that a basic juxtamembrane region corresponds to a DNA-binding motif essential for the activation of YabT trans-autophosphorylation. YabT is expressed during spore development and localizes to the asymmetric septum where it specifically phosphorylates essential proteins involved in genome maintenance, such as RecA, SsbA, and YabA. YabT has also been shown to phosphorylate proteins involved in protein synthesis, such as AbrB and Ef-Tu, suggesting a possible regulatory role in the progressive metabolic quiescence of the forespore. Finally, cross phosphorylations with other protein kinases implicate YabT in the regulation of numerous other cellular processes. Using an artificial protein scaffold as crystallization helper, we determined the first crystal structure of this DNA-dependent bacterial protein kinase. This allowed us to trap the active conformation of the kinase domain of YabT. Using NMR, we showed that the basic juxtamembrane region of YabT is disordered in the absence of DNA in solution, just like it is in the crystal, and that it is stabilized upon DNA binding. In comparison with its closest structural homolog, the mycobacterial kinase PknB allowed us to discuss the dimerization mode of YabT. Together with phosphorylation assays and DNA-binding experiments, this structural analysis helped us to gain new insights into the regulatory activation mechanism of YabT.
KW - Autophosphorylation
KW - Dimerization
KW - Regulatory mechanism
KW - Crystallization chaperone
KW - Spore development
U2 - 10.3389/fmicb.2018.03014
DO - 10.3389/fmicb.2018.03014
M3 - Journal article
C2 - 30671027
SN - 1664-302X
VL - 9
JO - Frontiers in Microbiology
JF - Frontiers in Microbiology
M1 - 3014
ER -