Specification of Hsp70 Function by Type I and Type II Hsp40 - The Networking of Chaperones by Co-chaperones - page 95

Sis1 ∆G/F still binds denatured luciferase and the RNQ1 protein, and Sis1 ∆G/F
still stimulates the Hsp70 ATPase activity. The function that was lacking in the Sis1
∆G/F protein was the ability to cooperate with Hsp70 to refold denatured substrates
(Aron et al. 2005 , 2007 ; Sondheimer et al. 2001 ; Johnson and Craig 2001 ). Sis1
∆G/F can still bind substrates and stimulate ATPase activity, so the defect likely
comes from an inability to efficiently transfer substrates from Sis1 to Hsp70. In
support of this conclusion mutation of the conserved ASP-ILE-PHE (DIF) motif
in the G/F region interferes with functions of Hsp40s that occur after J-domain
dependent hydrolysis of ATP by Hsp70 (Cajo et al. 2006 ). Molecular details of G/F
region action in Hsp40 function require further study (Wall et al. 1995 ), and this is
an important topic because this domain clearly plays a critical role in regulation of
Hsp70 function.
Central Domains
In addition to the differences found in the G/F regions, the central domains of the
Type I and Type II Hsp40s also have dramatic structural differences(Borges et al.
2012 ; Silva et al. 2011 ; Ramos et al. 2008 ). The central domain of the Type II
Hsp40s contains the G/M region and a polypeptide-binding site found in CTD1,
while the Type I Hsp40s contain a ZFLR that is adjacent to CTDI. The differences
in the substrate binding domains will be discussed in the next section, so for now we
will concentrate on how the G/M region versus the ZFLR may help specify func-
tion. Studies with the full length Sis1 protein indicate that the G/M region has some
overlapping function with the G/F region. As discussed above, deletion of unique
residues within the G/F region has deleterious effects on cell growth in cells that
only have a truncated version of Sis1 containing the J domain and G/F region (Aron
et al. 2005 ; Johnson and Craig 2001 ). However, in cells expressing the full length
Sis1, deletion of the same unique residues, Sis1 ∆101-113, no longer effects cell
growth at normal temperatures. These cells also maintain the prion state of RNQ1.
Likewise, deletion of the G/M region from the full-length protein (Sis 1 ∆G/M) has
no effect on cell growth at normal temperatures and has a very mild effect on the
maintenance of the RNQ1 prion. However, deletion of both the G/M and the unique
residues within the G/F region from the full-length protein (Sis 1 ∆G/M ∆101-113)
prevents the maintenance of the RNQ1 prion. These studies indicate that the es-
sential function of Sis1 is actually specified by both the G/M region and the unique
residues within the G/F region (Aron et al. 2005 ; Johnson and Craig 2001 ).
Studies of the ZFLR of Type I Hsp40s has also provided clues as to why the
function of the Type I proteins is unique from the Type II proteins. While the cen-
tral domain of the Type I Hsp40s, the ZFLR, has been implicated as a component
of the polypeptide binding site in combination with CTDI. The exact role of the
ZFLR is not completely clear. A NMR structure of the ZFLR reveals a V-shaped
groove with an extended B-hairpin topology, which could potentially be involved in
protein:protein interactions (Martinez-Yamout et al. 2000 ). A proteolytic fragment
of Ydj1, Ydj1 (179-384), which is missing the J-domain and the first zinc binding