Intermediates in the guanine nucleotide exchange reaction of Rab8 protein catalyzed by guanine nucleotide exchange factors Rabin8 and GRAB.
Small G-proteins of the Ras superfamily control the temporal and spatial coordination of intracellular signaling networks by acting as molecular on/off switches. Guanine nucleotide exchange factors (GEFs) regulate the activation of these G-proteins through catalytic replacement of GDP by GTP. During nucleotide exchange, three distinct substrate·enzyme complexes occur: a ternary complex with GDP at the start of the reaction (G-protein·GEF·GDP), an intermediary nucleotide-free binary complex (G-protein·GEF), and a ternary GTP complex after productive G-protein activation (G-protein·GEF·GTP). Here, we show structural snapshots of the full nucleotide exchange reaction sequence together with the G-protein substrates and products using Rabin8/GRAB (GEF) and Rab8 (G-protein) as a model system. Together with a thorough enzymatic characterization, our data provide a detailed view into the mechanism of Rabin8/GRAB-mediated nucleotide exchange.
From the Department of Physical Biochemistry, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany.
Zhong
Z
Guo
From the Department of Physical Biochemistry, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany,; the College of Life Science, Qingdao Agricultural University, Qingdao 266109, China.
Xiaomin
X
Hou
From the Department of Physical Biochemistry, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany,. Electronic address: roger.goody@mpi-dortmund.mpg.de.
Roger S
RS
Goody
From the Department of Physical Biochemistry, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany,; the Center for Integrated Protein Science Munich, Chemistry Department, Technische Universität München, 85747 Garching, Germany. Electronic address: aymelt.itzen@tum.de.
Aymelt
A
Itzen
D002384:Catalysis; D000081410:Germinal Center Kinases; D006160:Guanosine Triphosphate / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D009097:Multienzyme Complexes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020826:rab3A GTP-Binding Protein / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D000081410:Germinal Center Kinases; D009097:Multienzyme Complexes; D011994:Recombinant Proteins; D006160:Guanosine Triphosphate; D017346:Protein Serine-Threonine Kinases; C082377:RAB8A protein, human; D020691:rab GTP-Binding Proteins; D020826:rab3A GTP-Binding Protein
Enzyme Mechanisms; Guanine Nucleotide Exchange Factor (GEF); Intracellular Trafficking; Rab Proteins; Small GTPases