The structural differences of five forms of alpha-lactalbumin as revealed by FTIR spectroscopy.
Item
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Title
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The structural differences of five forms of alpha-lactalbumin as revealed by FTIR spectroscopy.
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Identifier
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AAI9924860
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identifier
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9924860
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Creator
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Zhong, Hui.
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Contributor
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Adviser: Robert Callender
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Date
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1999
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Language
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English
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Publisher
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City University of New York.
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Subject
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Chemistry, Biochemistry | Biophysics, General
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Abstract
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We have performed heat induced thermal denaturation study on bovine a -lactalbumin by application of Fourier transformation spectroscopy. Different modified forms of this protein, including the apo form, the acid form, the cam-3ss form and the apo cam-3ss form, were studied in addition to the intact so as to find the influence of the ligand calcium and disulfide bond 6-120 on the folding and stability of a -lactalbumin. The transition curves of the intact protein indicate gradual melting behavior of the b -structure below 60 ° C. The major transition happening in the mid 60 ° C involves the denaturation of helical structures. No native b -structure is found in the acid form. The cam-3ss form exhibits similar melting to the intact protein. However, the major transition temperature shifts down to the mid fifties as result of the broken disulfide bond. Therefore, a major role of the disulfide bond 6-120 is to stabilize protein. The transition curves of the apo form of a -lactabumin show two transitions instead of one transition found in the calcium bound forms. The transition at low temperature (around 20 ° C) involves substantial decrease of the band at 1638 and 1653 cm -1, indicative of the denaturation of the helical structures. On the other hand, the band at 1629 cm-1 increases, suggesting the increased formation of b -structure. The second transition happens around 40°C. The remaining components, largely b -structure, are unfolded through this transition. The apo cam-3ss form of a -LA shows one major transition in the experimental range. Further analysis of the spectra suggests a low temperature transition below 5 ° C involving melting of the helical structures. The high temperature transition happens around 40 ° C and is related to the unfolding of the b -structure, similar to that of the apo form. The non-cooperative transition behavior of the two structural domains in the calcium free forms of a -lactalbumin suggests these two domains melt independently. This thermal denaturation study further infers that b -domain folds first, followed by binding of calcium ion, which then triggers the complete folding of the helical domain during the folding process of a -lactalbumin from molten globule to native protein.
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
