and its primary structure was found to be 40% iden- 
tical to that of a protein of the eukaryotic cytosol, 
t-complex polypeptide- 1 (TCPl). 
Thermophilic factor 55 (TF55 ) has quaternary 
structure of a chaperonin. TF55 was purified from 
Sulfolobus shibatae, a thermophilic archaebacter- 
ium that normally grows at 75°C. TF55 is a major 
55-kDa polypeptide in S. shibatae at this tempera- 
ture, but after a shift to the near-lethal temperature 
88°C, it is virtually the only protein synthesized. Its 
production at 88°C correlates with acquisition of 
thermotolerance: cells incubated first at 88 °C can 
survive subsequent shift to the otherwise lethal tem- 
perature of 95°C. 
In sucrose density-gradient sedimentation, TF55 
migrated as a 20S homo-oligomer, a behavior remi- 
niscent of the chaperonins. In scanning transmission 
electron microscopy, carried out in collaboration 
with Dr. Joseph Wall, a double-ring structure was 
observed, resembling that of the chaperonins. In- 
stead of seven-member rings, however, TF55 com- 
plex contained nine-member rings. 
Purified TF55 complex has biochemical proper- 
ties of a molecular chaperone. In collaboration 
with Dr. Hartl, Dr. Horwich and his colleagues ob- 
served that TF55 complex binds mesophilic pro- 
teins as they become thermally unfolded at 56°C 
and 70°C but does not bind thermophilic proteins 
that remain in native form at these temperatures. 
Like the chaperonins, TF55 complex exhibits 
ATPase activity, with the rate of hydrolysis at 75°C 
equivalent to that of groEL at 37°C. 
Predicted amino acid sequence of TF55 is 40% 
identical to that of a eukaryotic protein, TCPl. 
TCPl was originally identified as an abundant 58- 
kDa protein in mouse testis that mapped to the t- 
locus. A role in male-specific transmission ratio dis- 
tortion was proposed, but the protein subsequently 
proved to be ubiquitous in mammalian tissues and 
to be present also in Drosophila and Saccharo- 
myces cerevisiae. In yeast the gene was shown to be 
essential, and a cold-sensitive mutant exhibited an 
abnormal-appearing mitotic spindle. 
TCPl complex is a molecular chaperone in tu- 
bulin biogenesis. In collaboration with Dr. Himan 
Sternlicht and his co-workers, a role for TCPl as a 
chaperone in the biogenesis of tubulin was assessed. 
Dr. Sternlicht's group had previously observed that 
both a- and jS-tubulin subunits translated in reticulo- 
cyte lysate were transiently associated with a 900- 
kDa complex prior to appearance in monomer or 
a-/3 heterodimer forms. Release of newly translated 
tubulin subunits from the 900-kDa complex could 
be blocked by depletion of ATP with apyrase. Tubu- 
lin subunits associated with the complex were 
found to be exquisitely protease sensitive, com- 
pared with a high degree of protease resistance of 
monomer or heterodimer forms corresponding to 
that of native tubulin. This suggested that while 
bound by the complex, tubulin was present in an 
unfolded conformation, but in the presence of 
MgATP it was folded to a native conformation. 
Analysis of the purified 900-kDa complex re- 
vealed a collective of at least seven species of appar- 
ent molecular size 55-60 kDa. One of these species, 
a 58-kDa protein, gave a signal on immunoblot anal- 
ysis with a monoclonal antibody against mouse 
TCPl. The TCPl -containing complex thus differs 
from the chaperonins insofar as it appears to be a 
hetero-oligomer. How general its role maybe in me- 
diating protein folding in the eukaryotic cytosol re- 
mains to be determined by in vivo studies of both a 
temperature-sensitive mutant affecting yeast TCPl, 
isolated in Dr. Horwich's laboratory, and mutants 
affecting the other members of the complex. 
Dr. Horwich is also Associate Professor of Genet- 
ics and of Pediatrics at Yale University School of 
Medicine. 
Books and Chapters of Books 
Horwich, A.L., Hartl, F.-U., and Cheng, M.Y. 
1991. Role of hsp60 in folding mitochondrial 
proteins. In Heat Shock (Maresca, B., and Lind- 
quist, S., Eds.). New York: Springer- Verlag, pp 
165-173. 
Articles 
Koll, H., Guiard, B., Rassow, J., Ostermann, J., Hor- 
wich, A.L., Neupert, W., and Hartl, F.-U. 1992. 
Antifolding activity of hsp60 couples protein im- 
port into the mitochondrial matrix with export to 
the intermembrane space. Cell 68:1 163-1 175. 
Trent, J.D., Nimmesgern, E., Wall, J. S., Hartl, F.-U., 
and Horwich, A.L. 1 99 1 . A molecular chaperone 
from a thermophilic archaebacterium is related to 
the eukaryotic protein t-complex polypeptide- 1 . 
Nature 354:490-493. 
Wienhues, U., Becker, K., Schleyer, M., Guiard, B., 
Tropschug, M., Horwich, A.L., Pfanner, N., and 
Neupert, W. 1991. Protein folding causes arrest 
of preprotein translocation into mitochondria in 
vivo, f Cell Biol 115:1601-1609. 
Yaffe, M B., Farr, G.W., Miklos, D., Horwich, A.L., 
Sternlicht, M.L., and Sternlicht, H. 1992. TCPl 
complex is a molecular chaperone in tubulin bio- 
genesis. Nature 358:245-248. 
GENETICS 203 
