We have identified a novel human cDNA with a predicted protein sequence that has 28% amino acid identity with the E. coli Hsp70 cochaperone
GrpE and designated it HMGE. Even with this low level of amino acid identity the human sequence could be efficiently modelled
on the X-ray structure of the E. coli protein, suggesting that there may be significant functional conservation. Indeed, HMGE expressed in E.
coli as a GST fusion protein co-purified with the E. coli Hsp70 protein DnaK in the absence of ATP. DnaK could be released from the GSTHMGE
with a Mg-ATP wash. Subcellular fractionation and immunocytochemistry studies using antisera raized against HMGE show that it
is a mitochondrial protein. In contrast to studies of rat GrpE, however, HMGE also appears to bind the constitutive cytosolic Hsp70, Hsc70, in
addition to mitochondrial Hsp70, Mt-Hsp70. We have previously shown that Hsc70 nucleotide-exchange is rate limiting in the presence of
the DnaJ-protein, HSJ1b. However, HMGE was found to inhibit the HSJ1b-enhanced Hsc70 ATPase activity and may mediate this inhibition
by binding the DnaJ-protein, HSJ1b. This is the first description of a direct interaction between a DnaJ protein and GrpE-like protein. These
studies suggest that the structure of GrpE has been conserved throughout evolution and that the conserved structure can interact with several
forms of Hsp70, but that HMGE cannot form part of the reaction cycle for cytosolic Hsc70. q 2001 Elsevier Science B.V. All rights
reserved.