Berson, Olga (1996) The study of copper bioavailability and mechanism of uptake in the type I methanotroph Methylomicrobium albus BG8. Dissertation (Ph.D.), California Institute of Technology. http://resolver.caltech.edu/CaltechETD:etd-02062008-105513
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Two aspects of copper uptake by the type I methanotroph Methylomicrobium albus BG8 were investigated — the effect of copper speciation in the growth medium on copper accumulation and mechanisms of copper transport in this microorganism.
Copper accumulation in M. albus BG8 consisted of nonspecific sorption of copper to outer cell layers and copper internalization. Most of the copper accumulated by the cells was nonspecifically sorbed to the cellular surface and was removable by EDTA. This phenomenon was especially prominent when cultures were grown at high total copper concentrations in the growth medium. This reversible binding of copper to external sites (e.g., amino acid, carboxylic, hydroxy groups, etc.) was described by a hyperbolic model with a mean maximum binding capacity of [...] moles/cell and an apparent half saturation constant of [...] moles/l. Copper availability to M. albus BG8 was related to the cupric ion concentration rather than to that of total copper added to the growth medium. Total internalized copper (total copper not removable by EDTA) was relatively constant at [...] moles of copper per cell despite a 100—fold variation in medium total copper and cupric ion concentrations, indicating the presence of a specific homeostasis mechanism for copper.
A specific copper uptake system is expected to be copper-regulated. Several copper-regulated polypeptides were identified in both soluble and membrane cellular fractions. The corresponding gene of one copper-repressible polypeptide, corA, was cloned and sequenced. CorA appeared to be vital for M. albus BG8 since an insertion mutant defective in the gene grew very poorly on plates or in liquid culture. It was suggested that CorA might be a divalent metal porin.
Three putative copper ATPase genes, atpA, atpB, atpC, were cloned and the complete sequence of atpA was obtained. The gene product of atpA contained a copper-binding signature motif, suggesting that AtpA does play a role in copper transport in M. albus BG8.
A hypothesis for copper uptake in M. albus BG8 was suggested by this research. A copper-repressible protein (CorA) described in Chapter Three may be an outer membrane porin that works in tandem with the putative copper ATPase(s) (Chapter Four). Such a porin would not necessarily be specific for copper, and might be overexpressed under conditions of any divalent metal limitation to facilitate the metal diffusion into periplasm.
|Item Type:||Thesis (Dissertation (Ph.D.))|
|Degree Grantor:||California Institute of Technology|
|Division:||Engineering and Applied Science|
|Major Option:||Environmental Science and Engineering|
|Thesis Availability:||Restricted to Caltech community only|
|Defense Date:||30 May 1996|
|Default Usage Policy:||No commercial reproduction, distribution, display or performance rights in this work are provided.|
|Deposited By:||Imported from ETD-db|
|Deposited On:||29 Feb 2008|
|Last Modified:||26 Dec 2012 02:30|
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