The crotoxin complex, a high resolution electron microscopy study.

Persistent Link:
http://hdl.handle.net/10150/184439
Title:
The crotoxin complex, a high resolution electron microscopy study.
Author:
Degn, Laura Lee
Issue Date:
1988
Publisher:
The University of Arizona.
Rights:
Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
Abstract:
The crotoxin complex protein is the major neurotoxic component in the venom of the Brazilian rattlesnake, Crotalus durissus terrificus. The purified protein can be crystallized in the form of thin platelets (less than 500 Å thick) suitable for electron crystallography and image processing techniques. The unit cell dimensions of the crystal are a = 38.8 Å, b = 38.8 Å, c = 256 Å, and α = β = γ = 90°. These crystals can grow in layers. For a three-dimensional image reconstruction this necessitates the determination of the crystal thickness in order to combine information from low dose images of crystals of the same thickness. In the past, the highest resolution image (to 3.9 Å) recorded from a crotoxin complex crystal on the electron microscope was from a crystal embedded in glucose. However, since glucose cannot be removed in order to accurately determine the crystal thickness, a different embedding technique (amorphous ice embedding) and tried. It was determined that high resolution image information (to 3.9 Å) can be recorded from crotoxin complex crystals embedded in amorphous ice. Five images of crystals preserved in amorphous ice, all exhibiting resolution to at least 9 Å, were first processed by a global averaging method from which two-dimensional projection maps were calculated. These maps were not interpretable due to variations in the images as demonstrated by a second processing method. In the second processing method the images were divided into smaller areas, or patches, and these patches were averaged. The patchwork images produced from the second process indicate that there is variation across the original images. The most likely explanation for the variation is the bending, or lack of flatness, of the crystals on the grid. The determination of mass thickness of the crystals from optical density differences between the crystals and the carbon support in images of freeze-dried crystals was explored. It was found that this method could not determine the mass thickness of crotoxin complex crystals to within one layer (64 Å), but could clearly distinguish between one and three overlapping layers of freeze-dried purple membrane which was used as a test specimen.
Type:
text; Dissertation-Reproduction (electronic)
Keywords:
Rattlesnakes -- Venom.; Neurotoxic agents -- Chemical structure.; Electron microscopy.
Degree Name:
Ph.D.
Degree Level:
doctoral
Degree Program:
Molecular and Cellular Biology; Graduate College
Degree Grantor:
University of Arizona
Advisor:
Chiu, Wah

Full metadata record

DC FieldValue Language
dc.language.isoenen_US
dc.titleThe crotoxin complex, a high resolution electron microscopy study.en_US
dc.creatorDegn, Laura Leeen_US
dc.contributor.authorDegn, Laura Leeen_US
dc.date.issued1988en_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.description.abstractThe crotoxin complex protein is the major neurotoxic component in the venom of the Brazilian rattlesnake, Crotalus durissus terrificus. The purified protein can be crystallized in the form of thin platelets (less than 500 Å thick) suitable for electron crystallography and image processing techniques. The unit cell dimensions of the crystal are a = 38.8 Å, b = 38.8 Å, c = 256 Å, and α = β = γ = 90°. These crystals can grow in layers. For a three-dimensional image reconstruction this necessitates the determination of the crystal thickness in order to combine information from low dose images of crystals of the same thickness. In the past, the highest resolution image (to 3.9 Å) recorded from a crotoxin complex crystal on the electron microscope was from a crystal embedded in glucose. However, since glucose cannot be removed in order to accurately determine the crystal thickness, a different embedding technique (amorphous ice embedding) and tried. It was determined that high resolution image information (to 3.9 Å) can be recorded from crotoxin complex crystals embedded in amorphous ice. Five images of crystals preserved in amorphous ice, all exhibiting resolution to at least 9 Å, were first processed by a global averaging method from which two-dimensional projection maps were calculated. These maps were not interpretable due to variations in the images as demonstrated by a second processing method. In the second processing method the images were divided into smaller areas, or patches, and these patches were averaged. The patchwork images produced from the second process indicate that there is variation across the original images. The most likely explanation for the variation is the bending, or lack of flatness, of the crystals on the grid. The determination of mass thickness of the crystals from optical density differences between the crystals and the carbon support in images of freeze-dried crystals was explored. It was found that this method could not determine the mass thickness of crotoxin complex crystals to within one layer (64 Å), but could clearly distinguish between one and three overlapping layers of freeze-dried purple membrane which was used as a test specimen.en_US
dc.description.noteDigitization note: two pages with number 50; pages are different; p. 52 missing from paper original and microfilm copy-
dc.typetexten_US
dc.typeDissertation-Reproduction (electronic)en_US
dc.subjectRattlesnakes -- Venom.en_US
dc.subjectNeurotoxic agents -- Chemical structure.en_US
dc.subjectElectron microscopy.en_US
thesis.degree.namePh.D.en_US
thesis.degree.leveldoctoralen_US
thesis.degree.disciplineMolecular and Cellular Biologyen_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.grantorUniversity of Arizonaen_US
dc.contributor.advisorChiu, Wahen_US
dc.contributor.committeememberDeatherage, Jamesen_US
dc.contributor.committeememberHewlett, Martinezen_US
dc.contributor.committeememberRayment, Ivanen_US
dc.contributor.committeememberWells, Michaelen_US
dc.identifier.proquest8820130en_US
dc.identifier.oclc701249857en_US
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