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| Protein Crystal Growth (PCG) |
| 101.3 |
| Prime: Scott Parazynski |  |
| Backup: Pedro Duque | |
| Overview |
| One of the objectives of the PCG experiment is to grow and retrieve highly structured protein crystals that are large enough to be used to analyze the molecular structures of various proteins. The experiment is also designed to obtain information on the dynamics of protein crystallization so scientists can determine the parameters necessary to optimize the methods of producing large, high-quality, well-ordered crystals. In addition to using the microgravity of space to grow high-quality protein crystals for structural analysis, the experiments help develop technologies and methods to improve the protein crystallization process on Earth as well as in space. Vapor Diffusion PCG will grow crystals by the vapor diffusion method, which has been highly effective in previous shuttle experiments. In vapor diffusion, water evaporates from a protein solution and is absorbed by a more concentrated reservoir solution contained in a wicking material. As the protein concentration rises, the protein crystals form. The STES provides a stable growth environment by maintaining temperatures at 71.6 degrees Fahrenheit (22 degrees Celsius), plus or minus 0.1 degree Celsius. Single Locker Thermal Enclosure System (STES) The PCG single-locker thermal enclosure system (STES) payload consists of an STES, its contents, and various stowed items. The STES is a refrigeration and incubation module for conducting microgravity and biotechnology research in the orbiter crew compartment. It is installed in place of a shuttle middeck locker. PCG Hardware & Configurations PCG-STES can be flown in one of two possible configurations, designated as Block I and Block II. In the Block I configuration, STES contains a carrier that holds up to three second-generation vapor diffusion apparatus (VDA) trays. Each VDA contains 20 experiment chambers. The Block II configuration consists of a carrier containing protein crystallization apparatus for microgravity (PCAM) units. Each PCAM unit consists of multiple circular trays that contain seven chambers each. The trays are assembled in a stack, which forms a cylinder, and are activated simultaneously by a knob that is part of each PCAM unit. Each chamber consists of a reservoir that contains a wick injected with a concentrated precipitating agent and a sample well that contains the precipitant solution. On certain missions the customer may choose to fly hand-held diffusion test cell (HH-DTC) devices and/or ambient PCAM devices that are stowed separate from the STES. A single HH-DTC device consists of a column assembly containing four test cells in which proteins are diffused with precipitants and buffers. The four HH-DTC devices contain a total of 16 individual experiment test cells. The diffusion process is accomplished simultaneously in the four test cells of a single HH-DTC device by operation of a single experiment activation handwheel. Each HH-DTC device weighs approximately 2 pounds and has an approximate stowage volume of 12 inches by 2.5 inches by 3 inches. Crew interface is required for removal of the HH-DTC devices from stowage and attachment in the mounting area, close-up video prior to and following activation, operation of the handwheels to initiate the crystal growth process, close-up video of each crystal growth cell at periodic intervals, and replacement of the units in their stowage locker. The HH-DTC units do not require orbiter power or any other orbiter interface. The ambient PCAM devices are identical to PCAM devices that are flown in the STES as the Block II configuration. These PCAMs do not require active thermal control and are flown under ambient conditions in a standard middeck stowage locker. The ambient PCAMs remain in the locker during all mission phases except activation and deactivation. The original PCG vapor diffusion apparatus was used for more than 20 shuttle experiments and produced highly ordered crystals for analysis on Earth. The second-generation apparatus, which is making its fourth flight, is designed to improve the mixing of experiment solutions, especially certain solutions that are too viscous to be mixed adequately in the original hardware. Better mixing is expected to result in the formation of larger, higher quality protein crystals. The new design introduces a triple-barreled syringe, which replaces the double-barreled syringe of the original apparatus, to improve mixing of experiment solutions. The temperature of the STES is controlled by conducting heat in or out of its internal enclosure through the left wall. The heat transferred moves through a heat exchanger mounted on the left wall, which is heated or cooled by orbiter cabin air circulated by a fan in the unit. A maximum of 128 watts of nominal 28-volt dc electrical power is drawn from a single middeck outlet. The PCG-STES requires continuous power through all mission phases. Protein Handling and Preparation In general, purified proteins have a very short lifetime in solution; therefore, the PCG payload will be loaded on the shuttle no earlier than 24 hours before launch. Due to the instability of the protein crystals, the payload will be retrieved from the shuttle within three hours of landing. The payload will be battery-powered continuously from the time the samples are placed in the enclosures and loaded on the shuttle until it is recovered and delivered to the investigating team. For launch delays of more than 24 hours, the payload will need to be replenished with fresh samples. When the samples are returned to Earth, they will be analyzed by morphometry to determine size distribution and absolute/relative crystal size. They also will be analyzed with X-ray crystallography and biochemical assays of purity to determine their internal molecular order and protein homogeneity, respectively. The PCG payload is sponsored by NASA's Office of Life and Microgravity Sciences and Applications. The payload is developed and managed by the Center for Macromolecular Crystallography. |
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| History/Background |
| PCG investigations are conducted in space because space-grown crystals tend to be larger, purer, and more highly structured than Earth-grown crystals. Such crystals greatly facilitate the study of protein structures. Scientists want to learn about a protein's three-dimensional structure to understand how it works, how to reproduce it, or how to change it. X-ray crystallography is widely used to determine a protein's three-dimensional structure. This technique requires large, well-ordered crystals for analysis. During the past 12 years, several hardware configurations have been used to conduct protein crystal growth middeck experiments on space shuttle flights. The PCG payload is designed to conduct experiments that will supply information on the scientific methods and commercial potential for growing large high-quality protein crystals in microgravity. |
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| Benefits |
| Proteins are important, complex biochemicals that serve a variety of purposes in living organisms. Metabolic processes involving proteins play an essential role in our lives, from providing nourishment to fighting disease. In the past decade, rapid growth in protein pharmaceutical use has resulted in the successful application of proteins to insulin, interferons, human growth hormone, and tissue plasminogen activator. The pharmaceutical industry seeks these pure protein crystals because their purity will simplify Federal Drug Administration approval of new protein-based drugs. Pure, well-ordered protein crystals of uniform size are in demand as special formulations for use in drug delivery. Other potential applications include agricultural products and bioprocesses for use in manufacturing and waste management. Such research has attracted firms in the pharmaceutical, biotechnological, and chemical industries. In response, the Center for Macromolecular Crystallography (CMC), a NASA Center for the Commercial Development of Space at the University of Alabama in Birmingham, has formed affiliations with a variety of companies that are investing substantial amounts of time, research, and money developing protein samples for use in evaluating the benefits of microgravity. Structural information gained from protein crystal growth (PCG) activities can provide a better understanding of the body's immune system and aid in the design of safe and effective treatments for disease and infections. |
Editorial Contact Ed Campion
Technical Contact USA Web Master
