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From the
Historical Perspectives The pioneering work of the late Christian Anfinsen and
his colleagues (1) on the reoxidation of bovine pancreatic ribonuclease (RNase) to a native, biologically active enzyme in vitro
after reduction of disulfide bridges and disruption of tertiary
structure demonstrated that regeneration of native conformation of a
purified protein can occur spontaneously in a test tube without the
addition of any other co-factors or helper enzymes. This led to the
still valid conclusion that "no special genetic information, beyond that contained in the amino acid sequence, is required for the proper
folding of the molecule and for the formation of `correct' disulfide
bonds" (2).
Of course, the story of protein folding goes back much further
(reviewed in Ref. 3). A number of milestones can be noted. In 1911, Chick and Martin found that proteins could be denatured in
vitro, and they distinguished that process from aggregation of the
protein. In 1929, Wu postulated that protein denaturation was an
unfolding process and that native protein structures involved regular,
repeated patterns of folding into a three-dimensional network. Anson
and Mirsky in 1931 and Anson (1945) showed that hemoglobin folding is
reversible and that hemoglobin could be renatured in vitro
to a form that had a native-like absorption spectrum, oxygen binding,
and tryptic digestion pattern. Studies in the 1950s by Eisenberg and
Schwert and by Schellman demonstrated that denaturation and
renaturation are thermodynamic processes, involving a change in free
energy and large changes in conformation between the denatured and
native states.
Even the early investigators realized that the protein folding
processes that occurred in test tubes, although they could reconstitute
native structure, were too slow to work inside cells. For example, even
under optimized conditions of protein dilution, pH, and temperature,
renaturation of RNase takes about 20 min (2), and RNase is a relatively
simple monomeric protein. Renaturation of some multidomain proteins may
take several hours in vitro, yet it is clear that all
possible conformations could not be sampled on the way to native
structure. Levinthal (4) summed this up succinctly in the "Levinthal
paradox" that can be stated as follows: if a given amino acid can
assume approximately 10 different conformations, the total number of
possible conformations in a polypeptide chain of 100 residues would be
10100. The time that this could take would be well beyond
the life span of an organism if not of the universe, depending on how
many conformations could be sampled before a protein reaches native state. Thus, it was realized early on that cells must have special ways
to make the process more efficient.
Experiments to examine the role of the intracellular environment in
protein folding involved the renaturation of proteins such as RNase
(2), bovine pancreatic trypsin inhibitor
(BPTI)1 (5), or influenza hemagglutinin (6)
in isolated microsomal fractions. The results indicated that protein
folding can be facilitated by proteins contained in the endoplasmic
reticulum (ER) of eukaryotic cells. In the case of disulfide
bond-containing proteins such as BPTI (5) or the human chorionic
gonadotropin (hCG)- It was soon realized that many polypeptides can reform native structure
easily by themselves in vitro (usually small single domain
proteins) while others (more complex, multidomain, or oligomeric proteins) fold and assemble efficiently only in the presence of additional proteins that are not constituents of the final native protein itself. These additional proteins have been called "molecular chaperones."
The term molecular chaperone was first used by Laskey et al.
(8) to describe the role of nucleoplasmin in the assembly of DNA and
histones into nucleosomes. The name seemed appropriate because
nucleoplasmin promotes histone-histone interactions to form the correct
oligomeric form while preventing aggregation. It does so without itself
forming part of the nucleosome and without specifying nucleosome
structure. Hence nucleoplasmin assumes the role of a chaperone.
The term molecular chaperone has been applied by Ellis and Hemmingsen
(9) to the expanding families of proteins of bacterial and eukaryotic
compartments involved in protein folding, assembly, and translocation.
The term has stuck, and it is now used to define a wide variety of
factors that facilitate generation of native protein and nucleic acid
structures.
Protein Folding in Vitro Versus in Vivo There are some similarities as well as differences between
intracellular protein folding and protein folding in test tubes. For
instance, for the tailspike protein of Salmonella
typhimurium phage P22 (10, 11) and hCG- There is growing interest in what regulates the folding of mammalian
proteins in vivo because of the number of human diseases now
known to be related to protein folding defects (reviewed in Refs. 20
and 21). This includes cystic fibrosis, The intracellular folding pathway of only a few proteins has been
studied in detail. These include the S. typhimurium phage P22 tailspike protein (11), hCG-
Many of the eukaryotic proteins whose folding and assembly have been
studied in vivo are membrane or secreted proteins. They follow a similar route to the cell surface. (i) Synthesis is carried out in the rough ER. (ii) Nascent proteins are translocated into the
cisternal space of the ER where the signal peptide is cleaved; initial
co-translational folding involving secondary structure and some native
tertiary structure occurs; addition of high mannose N-linked
oligosaccharides and initial processing of N-linked
oligosaccharide chains (for glycoproteins) takes place; formation of
disulfide bonds occurs, and for multimeric proteins, oligomerization or subunit assembly is attained along with achievement of native structure. (iii) The proteins destined for the cell surface or secretion are translocated to the Golgi apparatus, further processed, and then either translocated to the cell surface or packaged into secretory vesicles for secretion.
Role of Disulfide Bond Formation in Protein Folding and
Assembly It has been clear for a long time that the in vitro
folding of proteins targeted for secretion is facilitated by folding in the presence of microsomal extracts (2). It is now known that microsomes contain many chaperones that foster protein folding (30) as
well as the systems to create a favorable redox potential for the
formation of disulfide bonds (13, 31). For many secreted proteins,
disulfide bonds are important for stabilization of tertiary structure
and for their assembly into multimeric structures (32). PDI is a key
factor facilitating disulfide bond-dependent folding. For
example, the rate of stimulation of BPTI folding by microsomal extracts
is enhanced to the extent expected by the PDI activity of these
extracts (5). Moreover, PDI stimulates the rate of folding of
kinetically trapped BPTI molecules in vitro by several thousand-fold but has little effect on disulfide bond formation of BPTI
molecules possessing disulfide bonds that form efficiently in the
absence of PDI (17).
The pioneering work of Creighton and his colleagues (reviewed in Ref.
34) employed the formation of intramolecular disulfide bonds as a
biochemical probe to study the folding pathway of BPTI. This 58-amino
acid protein has three intramolecular disulfide bonds, and when reduced
and denatured, its folding pathway can be followed in vitro
by the reformation of the disulfide bonds. Using alkylation with
iodoacetic acid to trap free thiols, Creighton et al. (34)
defined a folding pathway involving intermediates that varied in their
amount and type of disulfide bonds. They also observed the existence of
a significant, albeit mostly transient, population of intermediates
containing disulfide bonds not present in the native protein, the
formation of which is the result of disulfide bond rearrangements.
There is also evidence that disulfide bond rearrangement occurs during
protein folding in intact cells. For example, in the intracellular
folding pathway of the hCG- Another protein whose in vivo folding has been studied is
influenza hemagglutinin (HA) (28). The folding of HA in the ER has also
been followed by the formation of intrachain disulfide bonds. Folding
of HA starts cotranslationally with some disulfide bonds beginning to
form soon after both cysteines that are involved in a disulfide pair
enter the ER lumen. However, most disulfide bond formation in HA occurs
after nascent polypeptide bond termination. This has also been observed
during the in vivo folding of hCG- Role of Glycosylation in Protein Folding and Assembly Since many membrane and secretory proteins are glycoproteins, it
is important to consider the role of carbohydrates in protein folding,
assembly, and secretion. N-Linked oligosaccharides of the
high mannose composition are added cotranslationally to the Asn-X-Ser(Thr) consensus sequence of proteins in the ER. One
function of N-linked glycans is to facilitate protein
folding and conformational maturation. When N-linked chains
are eliminated by site-directed mutagenesis of Asn residues in
glycosylation consensus sequences or by treatment of cells with agents
that block addition of N-linked glycans or their processing,
many ER-synthesized glycoproteins misfold, aggregate, and get degraded
within the ER (reviewed in Ref. 40). Some glycoproteins seem to fold
and be translocated efficiently without their N-linked
glycans. The only rule that seems to emerge here is that larger, more
complex glycoproteins have more trouble folding if their
N-linked glycans are missing.
The role of N-linked oligosaccharide chains in intracellular
folding of the hCG- Molecular Chaperones The role of molecular chaperones in protein folding, assembly, and
intracellular translocation has been the subject of a number of recent
reviews (19, 42-45). ER chaperones play a key role in protein folding
and quality control. Cytosol chaperones play a key role in folding,
transport, and biological activity of a number of proteins targeted for
transport to specific organelles such as the nucleus and mitochondria.
Examples of some eukaryotic molecular chaperones are shown in Table
I.
Intracellular roles of eukaryotic protein chaperones (modified from
Gething and Sambrook (30))
Important members of the ER family of chaperones include BiP,
originally characterized as an immunoglobulin binding protein (hence
the name (46)), GRp (or ERp) 72 (47), GRp (or ERp) 94 (48), calnexin
(49), and calreticulin (50). Additional chaperones continue to be
discovered and characterized. The enzyme-like cofactors protein
disulfide isomerase (51) and peptidyl prolyl cis/trans-isomerase (52), which catalyzes the isomerization of trans- to cis-proline, are usually considered
as ER chaperones as well.
Chaperones appear to act sequentially in protein folding pathways by
binding to folding intermediates that are in various stages of folding
and then passing them on to the next chaperone or chaperone complex in
the cascade, eventually releasing a competent native protein (53-55).
Binding usually involves interaction of chaperones with hydrophobic
residues on the surface of unfolded proteins, and release often
involves ATP hydrolysis. Binding of chaperones does not involve
specific amino acid consensus sequences in the substrate protein but
rather is determined by the arrangement of hydrophobic residues.
Binding of BiP by folding intermediates, as an example, is favored by
7-8-residue amino acid sequences with aliphatic and aromatic amino
acid residues in alternating positions (56). These are the sort of
sequences that would normally be on the inside of native proteins,
providing a way for chaperones to discriminate between folded and
unfolded proteins.
What Happens to Misfolded Proteins? It is generally thought that misfolded proteins remain in the ER
and are sequestered and degraded there without being secreted. We now
know that there are some exceptions to this, e.g. certain mis(un)folded forms of the hCG- Mechanisms of Chaperone Action The observation that chaperones are needed to assist protein
folding in living cells does not negate the findings of Anfinsen and
others that proteins can fold spontaneously in solution based only on
information contained in their primary amino acid sequence. Indeed, the
data comparing the in vitro versus in vivo folding pathways
for proteins that have been studied in this regard, for example the
S. typhimurium phage P22 tailspike protein (10, 11) and the
hCG- The best evidence for the mechanisms by which chaperones assist protein
folding comes from Escherichia coli in which the chaperones DnaJ/DnaK and the chaperonins2 GroEL/GroES
act in concert to facilitate folding of proteins. Recent biochemical
evidence (reviewed in Refs. 19 and 45) and crystallographic data (58,
59) provide a fairly clear and fascinating story on this subject,
although there is still some controversy on some key points. GroEL is
made up of 14 identical 60-kDa subunits that form two heptameric ring
structures with a pocket in the middle that can accommodate proteins up
to about 60 kDa. GroES is a single heptameric ring structure of 10-kDa subunits that can form a cap over the GroEL structure and is involved in holding a folding intermediate within the GroEL pocket. Folding occurs in the GroEL pocket and involves binding and hydrolysis of ATP
and release and rebinding of incompletely folded protein within the
GroEL pocket until most of the protein is folded to native state
(19).
The data indicate that members of Hsp70 and Hsp40 families of
chaperones in E. coli, namely DnaK and DnaJ, respectively,
are involved in the initial binding of nascent polypeptides as they proceed off the ribosome. These chaperones assist in the early steps of
protein folding and can facilitate, in cooperation with the nucleotide
exchange factor GrpE, complete folding to a native state of some
proteins. Other proteins require the additional actions of the
GroEL/GroES system to complete folding (Fig. 2) (18).
Analogous systems exist in eukaryotic cells and most likely act through
mechanisms similar to the E. coli folding systems. The
cytosolic Hsc70 family and the Hsp70 ER analogue BiP appear to act like
the DnaK/DnaJ chaperones in E. coli. The TCP-1 family of
eukaryotic cystosolic chaperonins, of which the chaperonin TRiC is the
analogue of GroEL, appears to function like the GroEL/ES system. It
should be noted that not all proteins need to proceed through the above
noted Hsc70/Hsp70/Hsp40 and GroEL/GroES/TRiC systems to fold to native
structure. Genetic data using temperature-sensitive mutants to shut off
production of GroEL, for example, indicate that a maximum of about 30%
of E. coli proteins need GroEL to fold correctly (45).
Furthermore, four cellular folding compartments (the ER, mitochondrial
intermembrane space, and the chloroplast lumen of eukaryotic cells as
well as the periplasmic space of bacteria) do not contain GroEL-type
chaperonins. However, each of these compartments contains a complement
of chaperones that assist protein folding (Table I).
One key question that remains is whether proteins fold to native
structure while they are bound to chaperones or whether they have to be
released into solution to complete folding. Hartl and his colleagues
(53) support the former hypothesis. They have data to show that when
proteins are synthesized by a more in vivo-like translational system (reticulocyte lysate), sequential binding of
folding intermediates by Hsc70/Hsp40 and then by TRiC occurs and that
native state is achieved while a protein substrate remains bound to
TRiC. Lorimer and his colleagues (60), on the other hand, believe that
a polypeptide dissociates completely from GroEL during the folding
process and may rebind several times before it is folded correctly to
native state (so called "iterative annealing") but that final
folding events occur in solution. Whichever model turns out to be
correct, and it may be substrate-dependent, it is clear
that proteins folding at the high concentration and in the highly
compact state of the intracellular environment require chaperones in
order to assist their folding and prevent their spontaneous
aggregation.
FOOTNOTES REFERENCES
Volume 272, Number 6,
Issue of February 7, 1997
pp. 3125-3128
©1997 by The American Society for Biochemistry and Molecular Biology, Inc.
MINIREVIEW:
§¶ and§
Eppley Institute for Research in Cancer and
Allied Diseases, the § Department of Pharmacology, and the
¶ Department of Biochemistry and Molecular Biology, University of
Nebraska Medical Center, Omaha, Nebraska 68198-6805
Historical Perspectives
Protein Folding in Vitro Versus in Vivo
Role of Disulfide Bond Formation in Protein Folding and
Assembly
Role of Glycosylation in Protein Folding and Assembly
Molecular Chaperones
What Happens to Misfolded Proteins?
Mechanisms of Chaperone Action
FOOTNOTES
REFERENCES
subunit (7), the key ER protein involved appears
to be protein disulfide isomerase (see below).
subunit (7)
intermediates in the folding pathway of the proteins appear to be the
same in vivo and in vitro, but the rate and
efficiency with which proteins achieve final native state in
vivo is higher than that in vitro. It must also be kept
in mind that, both in vivo and in vitro, correct
folding is in competition with misfolding and aggregation. This depends
on the protein concentration used for in vitro folding reactions, and in general, very dilute protein concentrations (0.01-0.02 mg/ml) (2, 12) are needed to prevent aggregation. This has
presented a huge problem to the biotechnology industry in attempts to
produce useful amounts of recombinant proteins. The efficiency of
folding in vitro can frequently be facilitated by
appropriate adjustment of the redox potential (13-15) or the addition
of factors such as protein disulfide isomerase (PDI) for eukaryotic
disulfide-bonded proteins (15-17) or DnaK/DnaJ chaperones for
bacterial proteins (reviewed in Refs. 18 and 19). In contrast to what
happens in vitro, cells minimize or circumvent the
off-pathway events by utilizing molecular chaperones that facilitate
the folding process by preventing aggregation and other unfavorable
interactions.
1-antitrypsin deficiency, Alzheimer's disease, Creutzfeld-Jacob disease,
neurodegenerative diseases such as Huntington's chorea, and
cancer.
subunit (22-25), luciferase (26),
influenza hemagglutinin (27, 28), and the HIV type 1 envelope
glycoprotein (29). Where the pathways have been determined both
in vitro and in vivo, for example for the phage
P22 tailspike protein (10, 11) and the hCG- subunit (7, 22-25), the
in vitro and in vivo folding pathways proceed
through the same respective folding intermediates. A diagram of the
folding pathway of the hCG- subunit is shown in Fig.
1.
Fig. 1.
Model of the hCG- kinetic folding
pathway. Intracellular kinetic studies have indicated that newly
synthesized intracellular hCG- (p1-early) is converted
into a folding intermediate that has both the 34-88 and 38-57
disulfide bonds formed, p1-late (t1/2 = 2-3 min). Along with the sequential formation of the 9-90 and
23-72 disulfide bonds, p1-late undergoes a major conformational shift into p2-free (t1/2 = 4-5 min).
When the 93-100 disulfide bridge forms, p2-free is converted into
an assembly-competent intermediate (t1/2 = 8-10
min) that, following association with the subunit, is recognized as
early p2-combined. After heterodimer assembly occurs, the
26-110 bond forms a "seat belt" around the subunit. The
crystallographic data (38, 39) indicate that the disulfide bonds 38-90
and 9-57 are present in the mature hCG heterodimer, suggesting
that there is a disulfide rearrangement in the subunit folding
pathway (reprinted with permission from Bedows et al.
(36)).
[View Larger Version of this Image (19K GIF file)]
subunit determined by pulse-chase
kinetics (35) and by site-directed mutagenesis of cysteines involved in
disulfide bonds (36, 37), two of the six disulfide bonds formed during
the kinetic folding pathway of hCG- are different from those seen in
the crystal structure of the native protein (38, 39).
(35).
subunit has been determined by examining the
kinetics of folding in Chinese hamster ovary cells transfected with
wild-type or mutant hCG- genes lacking one or both of the asparagine
glycosylation sites (41). Folding of hCG- lacking both
N-linked glycans was inefficient and correlated with the slow formation of the last three disulfide bonds (i.e.
disulfides 23-72, 93-100, and 26-110) to form in the hCG- folding
pathway. Unglycosylated hCG- was slowly secreted from Chinese
hamster ovary cells, and subunit folding intermediates retained in
cells for more than 5 h were degraded into a smaller hCG-
fragment. However, coexpression of hCG-, which is required for
formation of the biologically active heterodimer, enhanced
folding and formation of disulfide bonds 23-72, 93-100, and 26-110
of hCG- lacking N-linked glycans, suggesting that the
presence of its heterodimeric companion subunit fosters subunit
folding and assembly, perhaps because the subunit can act like a
chaperone for subunit folding. In addition, the molecular
chaperones BiP, ERp72, and ERp94, were found in a stable complex with
unglycosylated, unfolded hCG- and may be involved in the folding of
this form (41). These data indicate that N-linked
oligosaccharides assist hCG- subunit folding by facilitating
disulfide bond formation, perhaps by increasing the stability and
solubility of the native structure that fosters disulfide
formation.
Organelle
Target polypeptide
Chaperone
Role
Mitochondria
Mitochondrial
precursors
Hsp70
Completion of translocation Stabilization
of prefolded structures in matrix
Precursors in
matrix
Hsp60
Stabilization of prefolded structures and folding
Re-export of precursors to intermembrane space
Endoplasmic reticulum
Nascent secretory proteins
BiP
(GRP78)
Completion of folding and translocation Stabilization of
prefolded structures in lumen
Unfolded
proteins
GRP72
Completion of folding
Unfolded
proteins
GRP94
Completion of folding
Unfolded
proteins
Calreticulin
Completion of folding (ER lumen)
Unfolded proteins
Calnexin (p88, IP90)
Completion of
folding (ER membrane bound)
Mutant or foreign
proteins
BiP
Stabilization of unfolded structures; target for
degradation
Subunits of T cell receptor
TRAP or
p28
Receptor assembly
MHC Class 1 heavy
chains
p88
Stabilization of newly synthesized heavy chains
Proline-containing proteins
cis/trans-Prolyl
isomerase
Catalysis of slow protein folding reactions
Disulfide bond- containing proteins
Protein disulfide
isomerase
Formation and rearrangement of disulfide bonds
Cytosol
Nascent polypeptides
Hsc70
Stabilization of
prefolded structures
Mitochondrial and secretory
precursors
Hsc70
Antifolding before translocation
Unfolded proteins
Hsp40
Functions in cooperation with
Hsc70 to stabilize non- native protein conformations
Clathrin-coated vesicles
Hsc70 (clathrin uncoating
ATPase)
Binds exposed loop of clathrin light chain to promote
uncoating
Aged proteins
Hsc70 (Prp73)
Targeting to
lysosomes for degradation
Steroid
receptors
Hsp90
Stabilizes inactive form of receptor
Hsp70
Stabilizes inactive receptor
Immunophilins (e.g. FKBP52 and -54;
cyclophilin-40)
Stabilizes inactive receptor
p23
Stabilizes inactive receptor
Retroviral
transforming proteins
Hsp90
Stabilizes inactive form of protein
during transit to plasma membrane
Unfolded/misfolded
proteins
TRiC
Binds protein folding intermediates and promotes
folding
Nucleus
Preribosomes
Hsp70/Hsc70
Protection of
heat denatured proteins
Histones
Nucleoplasmin
Nucleosome
assembly
subunit. Some mutant forms of hCG- that do not fold properly are degraded intracellularly while other mutant forms that remain incompletely folded are secreted (37).
Nevertheless, there are a number of examples where protein misfolding
leads to protein accumulation in the ER and degradation. Some molecular
chaperones appear to be involved in targeting irreversibly misfolded
proteins for degradation in the ER. BiP is one of these. For example,
immunoglobulin light chains that are slowly folding and retained in the
ER of cultured mouse cells are quantitatively bound to BiP as partially
disulfide-bonded forms and then degraded, whereas light chains that are
more rapidly folded and secreted only transiently interact with BiP
(57).
subunit (7), indicate that proteins go through the same folding
steps in vitro and intracellularly. What then do chaperones
do and why do cells need them?
Fig. 2.
Role of the Hsp70 machinery (DnaK, DnaJ, and
GrpE) and Hsp60 machinery (GroEL and GroES) in protein folding.
DnaK and DnaJ recognize and bind hydrophobic regions in short peptides, thus preventing premature folding or aggregation during translation and
translocation. The ATP-dependent binding and release of
peptides use GrpE for stimulating nucleotide exchange. Partially folded polypeptides may then be passed from the DnaK/DnaJ/GrpE machinery to
the GroEL/ES·Hsp60 complex of E. coli. This large
oligomeric complex allows completion of folding of some polypeptides
that require additional assistance to fold to native structure
(reprinted with permission from Wall and Plückthun (18)).
[View Larger Version of this Image (31K GIF file)]
1
The abbreviations used are: BPTI, bovine
pancreatic trypsin inhibitor; ER, endoplasmic reticulum; hCG, human
chorionic gonadotropin; PDI, protein disulfide isomerase; HA,
hemagglutinin.
2
The GroEL/GroES/TRiC family of folding factors
are called chaperonins. Functionally, they are members of the larger
family of chaperones. The chaperonins have in common the ability to
promote protein folding in their central cavity.
©1997 by The American Society for Biochemistry and Molecular Biology, Inc.
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 P. Béguin, U. Hasler, O. Staub, and K. Geering Endoplasmic Reticulum Quality Control of Oligomeric Membrane Proteins: Topogenic Determinants Involved in the Degradation of the Unassembled Na,K-ATPase alpha Subunit and in Its Stabilization by beta Subunit Assembly Mol. Biol. Cell, May 1, 2000; 11(5): 1657 - 1672. [Abstract] [Full Text]
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V. Singh and W. E. Merz Disulfide Bond Formation Is Not Required for Human Chorionic Gonadotropin Subunit Association. STUDIES WITH DITHIOTHREITOL IN JEG-3 CELLS J. Biol. Chem., April 14, 2000; 275(16): 11765 - 11770. [Abstract] [Full Text] [PDF]
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N. Bakalara, X. Santarelli, C. Davis, and T. Baltz Purification, Cloning, and Characterization of an Acidic Ectoprotein Phosphatase Differentially Expressed in the Infectious Bloodstream Form of Trypanosoma brucei J. Biol. Chem., March 17, 2000; 275(12): 8863 - 8871. [Abstract] [Full Text] [PDF]
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 L. GORZA and M. VITADELLO Reduced amount of the glucose-regulated protein GRP94 in skeletal myoblasts results in loss of fusion competence FASEB J, March 1, 2000; 14(3): 461 - 475. [Abstract] [Full Text]
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G. Giannoukos, A. M. Silverstein, W. B. Pratt, and S. S. Simons Jr. The Seven Amino Acids (547-553) of Rat Glucocorticoid Receptor Required for Steroid and Hsp90 Binding Contain a Functionally Independent LXXLL Motif That Is Critical for Steroid Binding J. Biol. Chem., December 17, 1999; 274(51): 36527 - 36536. [Abstract] [Full Text] [PDF]
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 N. K. Relan, Y. Yang, S. Beqaj, J. H. Miner, and L. Schuger Cell Elongation Induces Laminin {alpha}2 Chain Expression in Mouse Embryonic Mesenchymal Cells: Role in Visceral Myogenesis J. Cell Biol., December 13, 1999; 147(6): 1341 - 1350. [Abstract] [Full Text] [PDF]
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 A. J. L. Macario, M. Lange, B. K. Ahring, and E. C. De Macario Stress Genes and Proteins in the Archaea Microbiol. Mol. Biol. Rev., December 1, 1999; 63(4): 923 - 967. [Abstract] [Full Text] [PDF]
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 D. R. Pillai, P. S. K. Wan, Y. C. W. Yau, J. I. Ravdin, and K. C. Kain The Cysteine-Rich Region of the Entamoeba histolytica Adherence Lectin (170-Kilodalton Subunit) Is Sufficient for High-Affinity Gal/GalNAc-Specific Binding In Vitro Infect. Immun., August 1, 1999; 67(8): 3836 - 3841. [Abstract] [Full Text] [PDF]
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J. Bhattacharyya and K. P. Das Molecular Chaperone-like Properties of an Unfolded Protein, alpha s-Casein J. Biol. Chem., May 28, 1999; 274(22): 15505 - 15509. [Abstract] [Full Text] [PDF]
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 A. L. Fink Chaperone-Mediated Protein Folding Physiol Rev, April 1, 1999; 79(2): 425 - 449. [Abstract] [Full Text] [PDF]
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 I. Kurek, K. Aviezer, N. Erel, E. Herman, and A. Breiman The Wheat Peptidyl Prolyl cis-trans-Isomerase FKBP77 Is Heat Induced and Developmentally Regulated Plant Physiology, February 1, 1999; 119(2): 693 - 704. [Abstract] [Full Text]
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S. N. Sarkar, S. Bandyopadhyay, A. Ghosh, and G. C. Sen Enzymatic Characteristics of Recombinant Medium Isozyme of 2'-5' Oligoadenylate Synthetase J. Biol. Chem., January 15, 1999; 274(3): 1848 - 1855. [Abstract] [Full Text] [PDF]
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 J. F. Rippmann, M. Klein, C. Hoischen, B. Brocks, W. J. Rettig, J. Gumpert, K. Pfizenmaier, R. Mattes, and D. Moosmayer Procaryotic Expression of Single-Chain Variable-Fragment (scFv) Antibodies: Secretion in L-Form Cells of Proteus mirabilis Leads to Active Product and Overcomes the Limitations of Periplasmic Expression in Escherichia coli Appl. Envir. Microbiol., December 1, 1998; 64(12): 4862 - 4869. [Abstract] [Full Text]
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 D. F. Smith, L. Whitesell, and E. Katsanis Molecular Chaperones: Biology and Prospects for Pharmacological Intervention Pharmacol. Rev., December 1, 1998; 50(4): 493 - 514. [Abstract] [Full Text] [PDF]
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S. Guha, T. K. Manna, Kali. P. Das, and B. Bhattacharyya Chaperone-like Activity of Tubulin J. Biol. Chem., November 13, 1998; 273(46): 30077 - 30080. [Abstract] [Full Text] [PDF]
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U. Hasler, X. Wang, G. Crambert, P. Beguin, F. Jaisser, J.-D. Horisberger, and K. Geering Role of beta -Subunit Domains in the Assembly, Stable Expression, Intracellular Routing, and Functional Properties of Na,K-ATPase J. Biol. Chem., November 13, 1998; 273(46): 30826 - 30835. [Abstract] [Full Text] [PDF]
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F. D. Menozzi, R. Bischoff, E. Fort, M. J. Brennan, and C. Locht Molecular characterization of the mycobacterial heparin-binding hemagglutinin, a mycobacterial adhesin PNAS, October 13, 1998; 95(21): 12625 - 12630. [Abstract] [Full Text] [PDF]
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M. Muyan, R. W. Ruddon, S. E. Norton, I. Boime, and E. Bedows Dissociation of Early Folding Events from Assembly of the Human Lutropin {beta}-Subunit Mol. Endocrinol., October 1, 1998; 12(10): 1640 - 1649. [Abstract] [Full Text]
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V. Westphal, J. C. Spetzler, M. Meldal, U. Christensen, and J. R. Winther Kinetic Analysis of the Mechanism and Specificity of Protein-disulfide Isomerase Using Fluorescence-quenched Peptides J. Biol. Chem., September 25, 1998; 273(39): 24992 - 24999. [Abstract] [Full Text] [PDF]
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P. Beguin, U. Hasler, A. Beggah, J.-D. Horisberger, and K. Geering Membrane Integration of Na,K-ATPase alpha -Subunits and beta -Subunit Assembly J. Biol. Chem., September 18, 1998; 273(38): 24921 - 24931. [Abstract] [Full Text] [PDF]
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A.-P. Gimenez-Roqueplo, J. Celerier, G. Lucarelli, P. Corvol, and X. Jeunemaitre Role of N-Glycosylation in Human Angiotensinogen J. Biol. Chem., August 14, 1998; 273(33): 21232 - 21238. [Abstract] [Full Text] [PDF]
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M. Xu, K. D. Dittmar, G. Giannoukos, W. B. Pratt, and S. S. Simons Jr. Binding of hsp90 to the Glucocorticoid Receptor Requires a Specific 7-Amino Acid Sequence at the Amino Terminus of the Hormone-binding Domain J. Biol. Chem., May 29, 1998; 273(22): 13918 - 13924. [Abstract] [Full Text] [PDF]
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T. G. Rozell, D. P. Davis, Y. Chai, and D. L. Segaloff Association of Gonadotropin Receptor Precursors with the Protein Folding Chaperone Calnexin Endocrinology, April 1, 1998; 139(4): 1588 - 1593. [Abstract] [Full Text] [PDF]
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J. Pantel, P. Robert, F. Troalen, M. Kujas, D. Bellet, and J.-M. Bidart Characterization of Human Lutropin Carboxyl- Terminus Isoforms Endocrinology, February 1, 1998; 139(2): 527 - 533. [Abstract] [Full Text] [PDF]
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P. Niccoli, L. Fayadat, V. Panneels, J. Lanet, and J.-L. Franc Human Thyroperoxidase in Its Alternatively Spliced Form (TPO2) Is Enzymatically Inactive and Exhibits Changes in Intracellular Processing and Trafficking J. Biol. Chem., November 21, 1997; 272(47): 29487 - 29492. [Abstract] [Full Text] [PDF]
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T. Jin, Y. Gu, G. Zanusso, M. Sy, A. Kumar, M. Cohen, P. Gambetti, and N. Singh The Chaperone Protein BiP Binds to a Mutant Prion Protein and Mediates Its Degradation by the Proteasome J. Biol. Chem., December 1, 2000; 275(49): 38699 - 38704. [Abstract] [Full Text] [PDF]
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P. P. Li, A. Nakanishi, S. W. Clark, and H. Kasamatsu Formation of transitory intrachain and interchain disulfide bonds accompanies the folding and oligomerization of simian virus 40 Vp1 in the cytoplasm PNAS, February 5, 2002; 99(3): 1353 - 1358. [Abstract] [Full Text] [PDF]
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