Browsing by Author "Liou, Gunn-Guang"

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Identification of Protein Domains on Major Pilin MrkA That Affects the Mechanical Properties of Klebsiella pneumoniae Type 3 Fimbriae
(2012-05-15)
Chan, Chia-Han
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Chen, Feng-Jung
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Huang, Ying-Jung
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Chen, Shin-Yu
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Liu, Kuo-Liang
The Klebsiella pneumoniae type 3 fimbriae are mainly composed of MrkA pilins that assemble into a helixlike filament. This study determined the biomechanical properties of the fimbriae and analyzed 11 site-directed MrkA mutants to identify domains that are critical for the properties. Escherichia coli strains expressing type 3 fimbriae with an Ala substitution at either F34, V45, C87, G189, T196, or Y197 resulted in a significant reduction in biofilm formation. The E. coli strain expressing MrkAG189A remained capable of producing a normal number of fimbriae. Although F34A, V45A, T196A, and Y197A substitutions expressed on E. coli strains produced sparse quantities of fimbriae, no fimbriae were observed on the cells expressing MrkAC87A. Further investigations of the mechanical properties of the MrkAG189A fimbriae with optical tweezers revealed that, unlike the wild-type fimbriae, the uncoiling force for MrkAG189A fimbriae was not constant. The MrkAG189A fimbriae also exhibited a lower enthalpy in the differential scanning calorimetry analysis. Together, these findings indicate that the mutant fimbriae are less stable than the wildtype. This study has demonstrated that the C-terminal beta strands of MrkA are required for the assembly and structural stability of fimbriae.
Structural and Mechanical Properties of Klebsiella pneumoniae Type 3 Fimbriae
(2011-04-01)
Chen, Feng-Jung
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Chan, Chia-Han
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Huang, Ying-Jung
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Liu, Kuo-Liang
;
Peng, Hwei-Ling
This study investigated the structural and mechanical properties of Klebsiella pneumoniae type 3 fimbriae, which constitute a known virulence factor for the bacterium. Transmission electron microscopy and optical tweezers were used to understand the ability of the bacterium to survive flushes. An individual K. pneumoniae type 3 fimbria exhibited a helix-like structure with a pitch of 4.1 nm and a three-phase force-extension curve. The fimbria was first nonlinearly stretched with increasing force. Then, it started to uncoil and extended several micrometers at a fixed force of 66 +/- 4 pN (n +/- 22). Finally, the extension of the fimbria shifted to the third phase, with a characteristic force of 102 +/- 9 pN (n = 14) at the inflection point. Compared with the P fimbriae and type 1 fimbriae of uropathogenic Escherichia coli, K. pneumoniae type 3 fimbriae have a larger pitch in the helix-like structure and stronger uncoiling and characteristic forces.
Structure of native oligomeric Sprouty2 by electron microscopy and its property of electroconductivity
(2013-09-27)
Chen, Feng-Jung
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Lee, Kuan-Wei
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Lai, Chun-Chieh
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Lee, Sue-Ping
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Shen, Hsiao-Hsuian
Receptor tyrosine kinases (RTKs) regulate many cellular processes, and Sprouty2 (Spry2) is known as an important regulator of RTK signaling pathways. Therefore, it is worth investigating the properties of Spry2 in more detail. In this study, we found that Spry2 is able to self-assemble into oligomers with a high-affinity KD value of approximately 16 nM, as determined through BIAcore surface plasmon resonance analysis. The three-dimensional (3D) structure of Spry2 was resolved using an electron microscopy (EM) single-particle reconstruction approach, which revealed that Spry2 is donut-shaped with two lip-cover domains. Furthermore, the method of energy dispersive spectrum obtained through EM was analyzed to determine the elements carried by Spry2, and the results demonstrated that Spry2 is a silicon- and iron-containing protein. The silicon may contribute to the electroconductivity of Spry2, and this property exhibits a concentration-dependent feature. This study provides the first report of a silicon- and iron-containing protein, and its 3D structure may allow us (1) to study the potential mechanism through the signal transduction is controlled by switching the electronic transfer on or off and (2) to develop a new type of conductor or even semiconductor using biological or half-biological hybrid materials in the future. (C) 2013 Elsevier Inc. All rights reserved.