Bariloche protein symposium argentine society for biochemistry and molecular biology

34
BIOCELL, 27 (Suppl. I), 2003
S9.
METALLOPROTEIN DESIGN: ENGINEERING METAL-
BINDING SITES INTO NATIVE PROTEIN SCAFFOLDS
Lu, Yi
Department of Chemistry, University of Illinois at Urbana-
Champaign, Urbana, Illinois 61801, USA. E-mail: [e-mail]
Metalloproteins catalyze some of the most difficult biological
reactions and can fine-tune the reactivity at the highest level. While
much progress has been made on the study of native metalloproteins
and their variants, little is known about how to design a
metalloprotein with desired structure and activity. By using stable,
easy-to-produce, and well-characterized proteins as scaffolds, we
have successfully designed protein models of Cu
A
 and Cu
B
-heme
centers from cytochrome c oxidase, a Mn(II)-binding center from
manganese peroxidase and a heme-thiolate center from cytochrome
P450.
1
 Our successes provide an alternative approach to de novo
protein design. Nature is known to use only a limited number of
thermodynamically stable protein scaffolds and yet is able to
achieve diverse functions by designing different active sites into
the same scaffold. Therefore, as in nature, we can choose stable
scaffolds for designing metalloproteins. More importantly, the
study of the designed proteins has contributed to the understanding
of the structural and functional properties of several complex
metalloenzymes and has helped resolve long-standing issues in
the field. Recent progress in this endeavor will be presented.
1. Lu Y, Berry SM, Pfister TD (2001). Chem. Rev. 101: 3047-3080.
S10.
FROM SEQUENCE TO CONSEQUENCE
Ringe, Dagmar
Departments of Biochemistry and Chemistry and Rosenstiel Basic
Medical Sciences Research Center, Brandeis University  MS 029,
415 South Street, Waltham, MA 02454-9110, USA.
Genomics, the study of the properties of genes and gene products
on a whole-organism scale, is revolutionizing all aspects of biology.
The ultimate goals are the determinations of the functions of all
gene products of an organism within the context of the organism,
at all levels from the molecular to the organism as a whole.  Among
the fields that has the potential to aid in that process is the field of
structural biology.  Structural genomics has as its stated goals the
filling-in of the catalog of known protein folds and the assignment
of function to gene products whose functions are not known by
structural similarity to proteins of known function.  How realistic
are these goals?  How robust are the predictive methods that take
structure to function?  How can computational methods be used to
predict function?  These questions will be discussed in light of
the methods we have developed to address them.
S11.
STRUCTURAL PROPERTIES AND KINETIC ROLE OF
EARLY INTERMEDIATES IN PROTEIN FOLDING
Heinrich Roder, Hong Cheng, Ramil Latypov, Kosuke Maki and
M.C. Ramachandra Shastry.
Basic Science Division, Fox Chase Cancer Center, Philadelphia,
PA 19111. USA. E-mail: [e-mail]
While small globular proteins typically require milliseconds to
seconds to complete the process of folding, there is growing
evidence that important conformational changes, including
secondary structure formation and chain collapse, occur on a much
shorter time scale
1,2
. These early events are crucial for
understanding how protein folding is initiated and directed along
productive channels. A highly efficient capillary mixer
3
 coupled
with optical probes or NMR-detected H-D exchange has enabled
us to follow structure formation during folding over the time range
from tens of microseconds to minutes. Our observations of rapid
changes in fluorescence, absorbance and amide protection on the
sub-millisecond time scale provide insight into the structural
characteristics and kinetic role of early intermediates populated
during the folding of a diverse set of proteins, including cytochrome
c
4,5
, protein G
6
, Im7
7
, 
β-lactoglobulin
8
, staphylococcal nuclease
9
and acyl-CoA binding protein
10
.
1
Curr. Opin. Struct. Biol. 9, 620-626 (1999). 
2
Mechanisms of protein folding,
65-104 (Oxford University Press, New York, 2000). 
3
Biophys. J. 74, 2714-
2721 (1998). 
4
Nature Struct. Biol. 5, 385-392 (1998). 
5
J. Mol. Biol. 330,
1145-52 (2003). 
6
Nature Struct. Biol. 6, 943-947 (1999). 
7
Nature Struct.
Biol. 8, 68-72 (2001). 
8
Nature Struct. Biol. 8, 151-5. (2001). 
9
Protein Sci.
11, 82-91 (2002). 
10
Proc. Natl. Acad. Sci. USA 99, 9807-12 (2002).
S12.
HOW DO MUTANT COPPER-ZINC SUPEROXIDE
DISMUTASE PROTEINS KILL MOTOR NEURONS?
Valentine, Joan Selverstone
Department of Chemistry and Biochemistry, UCLA, Los Angeles,
California, USA. E-mail: [e-mail]
Amyotrophic lateral sclerosis (ALS ) is caused by selective death
of motor neurons. In a small fraction of the cases (familial ALS or
FALS), it is inherited as a autosomal dominant trait associated
with mutations in the gene encoding copper-zinc superoxide
dismutase (CuZnSOD). These mutations are known to confer new
and toxic properties on the protein and thereby cause the disease,
although the nature of the toxic property(ies) is still in dispute.
We have purified a series of ALS mutant human CuZnSODs and
found altered metal binding properties, chemical reactivities,
structures, and stabilities in the mutant proteins relative to the
wild type proteins. These and related observation may help to
explain what is perhaps the most perplexing question in CuZnSOD-
associated FALS, how such a diverse set of mutations could result
in the same gain of function that causes motor neurons to die.

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