3D Printing of Protein Models

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3D Printing of Protein Models Powered By Docstoc
					         3D Printing of Protein
                        Nick Stong
             University of Wisconsin-Madison

    George Phillips Ph.D. and Roman Aranda
 Departments of Biochemistry and Biostatistics and
               Medical Informatics
    College of Agricultural and Life Sciences
      School of Medicine and Public Health                                         One way to display the data in the PDB files is to ‘print’ a 3D
                                                                                   model. This is done using a Z Corporation printer. This lays
                                                                                   down a thin layer of plaster, and prints colored glue on the one
Introduction                                                                       layer, then lays down another layer of dust. This builds the
                                                                                   model layer by layer. This gives you a physical model that
The Protein Data Bank (PDB) was established in 1971 at the                         can be placed on a bookshelf. These representations of
Brookhaven National Laboratory; in 1998 it was transferred to                      proteins are important because it is the only way to get an
the Research Collaboratory for Structural Bioinformatics,                          actual 3D model, which is easiest to interpret and analyze. It
which is composed of Rutgers University, The University of                         is also very convenient since the model sits on a tabletop and
Wisconsin - Madison, NIST and the San Diego                                        is instantly accessible in a meeting, or can be passed around a
Supercomputer Center.          They maintain a variety of                          classroom. Proteins are prepared for printing by highlighting
information on 37,136 structures, all of which are made                            key interactions including, but not limited to, binding sites,
publicly available. Scientists all over the world using a variety                  domains, and other moving components of proteins. Protein
of techniques including x-ray crystallography and NMR                              models were prepared of Class I Major Histocompatabilty
spectroscopy determine this information experimentally. Most                       Complex (MHC) since it has an interesting binding site;
important is the 3-D model information available in .PDB files                     chaperronin because it is a complex protein whose mechanism
(Wikipedia).     Using Pymol, an open source molecular                             is difficult to picture; and myoglobin, since work was done on
visualization system, this data can be viewed and manipulated.                     it by Roman Aranda, a graduate student in George Phillips lab.
Using this program proteins can be explored and better
                                                                                   Class I MHC is part of the ‘antigen processing and
                                                                                   presentation’ pathway that takes portions of degraded proteins
                                                                                   from the cytosol and presents them on the cell surface for t-
                                                                                   cell recognition as native or foreign. The specific molecule
                                                                                   that was studied was the mouse H-2Dd MHC, complexed with
                                                                                   an immunodependant peptide derived from the human
                                                                                   immunodeficiency virus IIIB gp120 envelope glycoprotein,
                                                                                   P18-I10. This protein has a binding site nestled between the
                                                                                   a1 and a2 chains. The H-2Dd MHC uses a four-residue
                                                                                   binding motif made up of a glycine at P2, a proline at P3, a
                                                                                   positively charged residue at P5, and a C-terminal
                                                                                   hydrophobic residue at P9 or P10 (the oligopeptide is labeled
                                                                                   P1-P10)(figure 1). The other residues in the glycoprotein
                                                                                   bulge away from the binding pocket and are important for T-
                                                                                   Cell recognition (figure 1).

                                                                                   Chapperonin is a protein complex that chapperones proteins so
Figure 1. 3D model of Class I MHC. A1 and a2 chains are shown in green.
B Chain is shown in orange. HIV protein peptide residues important for T-
                                                                                   they will they fold into their native conformation. This is
Cell recognition; P2, P3, P5, P9, P10, are shown in pink. Other HIV peptide        done by identifying proteins that are beginning to fold
proteins are shown in purple.                                                      incorrectly, causing the protein to unfold slightly, and provide
                                                                                   a protected environment with more stable pH and protection

from negative interactions with other molecules. In E. coli                     monoxide ‘molecule’ within the center of the model(figure 3).
type I chapperonin complex consists of a GroEL cap and a                        This model CO trapped by physical spheres, however in the
GroEs cis and trans subunit. When a incorrectly folded                          protein the CO is trapped by molecular forces. This is the first
protein and 4 ATP molecules enter the complex the ATP binds                     model that is able to represent the movement within the
to the GroES cis subunit, causing a conformational change                       protein.
that binds the GroEL cap, creating a protected environment for
the protein to fold in. The model prepared shows the GroEL
cap and the the GroES cis subunit complexed with it. 4 bound
molecules of ADP and magnesium molecules important in the
binding of ADP are also shown (figure 2)

                                                                                Figure 3. 3D model of myoglobin. Heme group is shown in dark blue.
                                                                                Carbon monoxide molecule is trapped above heme group and ‘rattles’,
                                                                                showing movement.

Figure 2. 3D model of type I chapperonin. GroEL cap is shown in yellow.
GroES subunit is shown in green in cis conformation. ADP bound to induce
conformation change is shown in red

Myoglobin is a heme protein that reversibly binds small
gaseous ligands such as carbon monoxide (CO).; for this
reason myoglobin is found primarily in muscle and is
important in gas exchange. Myoglobin was one of the first
protein structures found and has been extensively studied. A                    Figure 4. Time Lapsed 3D of myoglobin. Time lapse structures are shown
new technique being applied to myoglobin is time resolved X-                    from red  purple. Movement shown is due to the forces exerted by the CO
ray crystallography, which provides a series of time lapsed                     molecule exiting from being bound to the heme
images of the protein at very brief intervals. By disrupting the
photoliable ligand-heme bond the dissociation of CO was                         Results
monitored over 100 ps to 3.16 ms. This provided 8 time
lapsed structures of hemoglobin, with the heme, CO and key                      The Class I MHC (Fig. 1) is colored to show the separate
residues shifted slightly in each structure. Roman Aranda did                   chains of the protein. The A chain is shown in green and the
this work in the Phillips lab. Color coding and overlapping of                  B chain is show in orange. Detail is only shown in the binding
these structures allows for the change over time of the protein                 site which is the area of interest in the protein. The protein is
to be seen in a physical representation (figure 2.). A                          bound to an HIV glycoprotein. The glycoprotein residues are
hemoglobin model was also made that traps a loose carbon                        colored individually in purple or pink. The proteins that are

important for binding and a part of the class I MHC binding
motif are shown in purple. The other residues are shown in
pink. Most of these residues are pushed away from the protein
and are important for T-cell recognition.

The chapperonin model (Fig. 2) is of a much larger protein. It
is easy to see how many individual residues are a part of the
model. The model shows how the chapperonin functions,
providing a large open space with controlled pH, ion
concentration, and charge that allows proteins to fold properly.

The first myoglobin model (Fig. 3) is meant to show
movement within the protein. Almost all representations of
proteins are static, when in fact they are dynamic molecules.
This model shows the movement of the separated carbon
monoxide within the heme pocket.

The second myoglobin model (Fig. 4) is also meant to show
movement of the carbon monoxide molecule as it is
dissociates from the myoglobin. This is done by overlapping
models at different time points. The models are colored
sequentially so one can follow the changes over time.

This work has given me the ability to be able to access and
intelligently interpret the data found in the protein data bank.
This skill has applications in any study at the molecular level
in which further understanding of the interaction between
proteins would be helpful. As 3D printing technology
becomes less complicated, and more automated, labs in nearly
all fields will be able to have physical representations of
studied structures. This will bring us one step further to fully
understanding the molecular world.

This research was supported by:
    • University of Wisconsin-Madison Graduate School
    • National Science Foundation (144 PE44)

1.   Li, H., et al. "Three-Dimensional Structure of H-2Dd
     Complexed with an Immunodominant Peptide from Human
     Immunodeficiency Virus Envelope Glycoprotein 120."
     Journal of Molecular Biology 283.1 (1998): 179-91.

2.   Roman Aranda IV, et al. "Time-Dependent Atomic
     Coordinates for the Dissociation of Carbon Monoxide from
     Myoglobin." International Union of Crystallography D.62
     (2006): 776-83.


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