 |
PDBsum entry 2v6y
|
|
|
|
References listed in PDB file
|
 |
|
Key reference
|
|
|
Title
|
|
Structural basis for selective recognition of escrt-Iii by the aaa atpase vps4.
|
|
|
Authors
|
|
T.Obita,
S.Saksena,
S.Ghazi-Tabatabai,
D.J.Gill,
O.Perisic,
S.D.Emr,
R.L.Williams.
|
|
|
Ref.
|
|
Nature, 2007,
449,
735-739.
[DOI no: ]
|
|
|
PubMed id
|
|
|
|
|
|
Abstract
|
|
|
The AAA+ ATPases are essential for various activities such as membrane
trafficking, organelle biogenesis, DNA replication, intracellular locomotion,
cytoskeletal remodelling, protein folding and proteolysis. The AAA ATPase Vps4,
which is central to endosomal traffic to lysosomes, retroviral budding and
cytokinesis, dissociates ESCRT complexes (the endosomal sorting complexes
required for transport) from membranes. Here we show that, of the six
ESCRT--related subunits in yeast, only Vps2 and Did2 bind the MIT (microtubule
interacting and transport) domain of Vps4, and that the carboxy-terminal 30
residues of the subunits are both necessary and sufficient for interaction. We
determined the crystal structure of the Vps2 C terminus in a complex with the
Vps4 MIT domain, explaining the basis for selective ESCRT-III recognition. MIT
helices alpha2 and alpha3 recognize a (D/E)xxLxxRLxxL(K/R) motif, and mutations
within this motif cause sorting defects in yeast. Our crystal structure of the
amino-terminal domain of an archaeal AAA ATPase of unknown function shows that
it is closely related to the MIT domain of Vps4. The archaeal ATPase interacts
with an archaeal ESCRT-III-like protein even though these organisms have no
endomembrane system, suggesting that the Vps4/ESCRT-III partnership is a relic
of a function that pre-dates the divergence of eukaryotes and Archaea.
|
|
|
|
|
|
|
|
Figure 1.
Figure 1: Characterization of the Vps4–Vps2 complex. a,
GST-tagged MIT domain of yeast Vps4 and full-length Vps4
interact with full-length Vps2. Coomassie-stained SDS–PAGE
shows the material bound to the glutathione-Sepharose resin. b,
c, Vps2 C-terminal constructs (residues 106–232 in b and
residues 120–232 in c) also interact with an untagged MIT
domain as detected by band-shift on native PAGE. d, Vps2
C-terminal region (residues 183–232) binds to the FlAsH-tagged
Vps4 MIT domain with a K[d] of 28  M.
e, Structure of the complex between Vps2 (cyan) and the Vps4 MIT
domain (yellow). f, Interactions between Vps2  C
and MIT domain. g, The distinctive three-corners-of-a-square
appearance of the three-helix MIT bundle. h–j, Enlarged
central (i) and peripheral Vps2 helix C
specificity determinants (N-terminal and C-terminal regions are
shown in h and j, respectively).
|
|
Figure 3.
Figure 3: MIT-interacting motifs in Vps2 and Did2 are important
for function in vivo. a, In wild-type cells, GFP-tagged
carboxypeptidase-S accumulates in the vacuolar lumen (FM4-64
preferentially labels the limiting membrane of the vacuole). A
single mutation at position 0 of the Vps2 MIM (R224D) impairs
sorting, and the GFP–CPS accumulates in the limiting
membrane of the vacuole. A double mutation of the Vps2 MIM
(L228D/K229D) causes GFP–CPS accumulation in a class E
compartment. b, Both a single mutation at position 0 of the Did2
MIM (R198D) and a double mutation (L199D/L202D) impair sorting
of GFP–CPS, which accumulates in the limiting membrane of the
vacuole.
|
|
|
|
|
|
The above figures are
reprinted
by permission from Macmillan Publishers Ltd:
Nature
(2007,
449,
735-739)
copyright 2007.
|
|
|
|
|
|
|
