Understanding the lytic domain of A2: the maturation protein of ssRNA bacteriophage QBeta

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Title: Understanding the lytic domain of A2: the maturation protein of ssRNA bacteriophage QBeta
Author: Langlais, Carrie-Lynn
Abstract: Most bacteriophage escape the confines of the host bacterium by compromising the integrity of its cell wall , an event that results in rupture (lysis ) of the cell . The lysis strategy of bacteriophage Q ? is inhibition of cell wall biosynthesis while the cell is growing . To elicit lysis , the maturation protein (A2 ) of Q ? inhibits the catalytic activity of MurA , an essential , induced fit enzyme in the cell wall biosynthetic pathway . Consequent lysis releases progeny phage into the environment . The research in this dissertation addresses how lysis timing is integrated into Q ? ?s life cycle and discerns the molecular basis of the lytic event . Working off the notion that , as displayed by the mature virion , A2 inhibits MurA , we developed an in vivo bioassay to resolve the amount of inhibitory A2 during infection . We found that the amount of free A2 is vastly greater than the amount of virion -associated A2 and that both forms inhibit MurA . Additionally , the amount of A2 correlates to lysis time and the burst size , as mutant Q ? with upregulated expression of A2 (Q ?por ) elicit host cell lysis faster and release fewer mature virions than with the wildtype level of A2 . This further suggests that protection from Q ? lysis afforded by MurAL138Q is due to perturbed affinity between A2 and MurA . Yeast two -hybrid analysis supports that A2 and MurAL138Q interact with weaker affinity by rendering small colonies compared to yeast containing interacting A2 -MurAwt . Scanning mutagenesis of MurA ?s surface near L138 identified residues that may be important for A2 contact in the inhibitory complex . Potentially important residues map to a contiguous area on the surface of MurA that spans both lobes on the flexible loop face of the enzyme , suggesting that A2 prevents the induced fit mechanism of MurA in an uncompetitive manner . Subsequent truncation analysis reveals that the aminoterminal half of A2 is sufficient to mediate host cell lysis . Together , these findings insinuate a model in which the amino -terminus of free A2 interacts with , and inhibits MurA . Then , when the infected cell initiates division , septal catastrophe ensues causing the cell to lyse and liberate progeny bacteriophage Q ? .
URI: http : / /hdl .handle .net /1969 .1 /ETD -TAMU -1239
Date: 2009-05-15

Citation

Understanding the lytic domain of A2: the maturation protein of ssRNA bacteriophage QBeta. Available electronically from http : / /hdl .handle .net /1969 .1 /ETD -TAMU -1239 .

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