Browsing by Author "Lin, Tiao-Yin"

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A T3 and T7 Recombinant Phage Acquires Efficient Adsorption and a Broader Host Range
(2012-02-09)
Lin, Tiao-Yin
;
Lo, Yi-Haw
;
Tseng, Pin-Wei
;
Chang, Shun-Fu
;
Lin, Yann-Tsyr
It is usually thought that bacteriophage T7 is female specific, while phage T3 can propagate on male and female Escherichia coli. We found that the growth patterns of phages T7M and T3 do not match the above characteristics, instead showing strain dependent male exclusion. Furthermore, a T3/7 hybrid phage exhibits a broader host range relative to that of T3, T7, as well as T7M, and is able to overcome the male exclusion. The T7M sequence closely resembles that of T3. T3/7 is essentially T3 based, but a DNA fragment containing part of the tail fiber gene 17 is replaced by the T7 sequence. T3 displays inferior adsorption to strains tested herein compared to T7. The T3 and T7 recombinant phage carries altered tail fibers and acquires better adsorption efficiency than T3. How phages T3 and T7 recombine was previously unclear. This study is the first to show that recombination can occur accurately within only 8 base-pair homology, where four-way junction structures are identified. Genomic recombination models based on endonuclease I cleavages at equivalent and nonequivalent sites followed by strand annealing are proposed. Retention of pseudo-palindromes can increase recombination frequency for reviving under stress.
iPARTS2: an improved tool for pairwise alignment of RNA tertiary structures, version 2
(2016-07-08)
Yang, Chung-Han
;
Shih, Cheng-Ting
;
Chen, Kun-Tze
;
Lee, Po-Han
;
Tsai, Ping-Han
Since its first release in 2010, iPARTS has become a valuable tool for globally or locally aligning two RNA 3D structures. It was implemented by a structural alphabet (SA)-based approach, which uses an SA of 23 letters to reduce RNA 3D structures into 1D sequences of SA letters and applies traditional sequence alignment to these SA-encoded sequences for determining their global or local similarity. In this version, we have re-implemented iPARTS into a new web server iPARTS2 by constructing a totally new SA, which consists of 92 elements with each carrying both information of base and backbone geometry for a representative nucleotide. This SA is significantly different from the one used in iPARTS, because the latter consists of only 23 elements with each carrying only the backbone geometry information of a representative nucleotide. Our experimental results have shown that iPARTS2 outperforms its previous version iPARTS and also achieves better accuracy than other popular tools, such as SARA, SETTER and RASS, in RNA alignment quality and function prediction. iPARTS2 takes as input two RNA 3D structures in the PDB format and outputs their global or local alignments with graphical display. iPARTS2 is now available online at http://genome.cs.nthu.edu.tw/iPARTS2/.
Protein-protein interaction as a powering source of oxidoreductive reactivity
(2010)
Lin, Tiao-Yin
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生物科技學系
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Department of Biological Science and Technology
Thiol-disulfide exchange reactions between thiol-disulfide oxidoreductases (e. g. thioredoxin or Trx) and client proteins can obtain a rate several orders faster than those between chemical reagents (e. g. dithiothreitol) and client proteins. The active sites of these oxidoreductases are characterized by a CXXC motif. The XX dipeptide of Trx is GP. By altering the C-terminal X to A, K and D, it is shown that the P -> K mutation confers the largest effect on the redox potential, which it elevated by 28 mV, while the P -> D mutation displays the smallest variation. The change in pK(a) of the nucleophilic thiol also follows this trend. However, GK and GA react faster with thioredoxin reductase, exhibiting a rate rank of GK > GA > GP > GD, while the rates toward insulin and PDI follow the order GP > GA > GK > GD. The rate change spans two to three orders of magnitude. This work demonstrates that redox reactivity does not correlate simply with pK(a) and redox potential, but instead supports the important role of interaction between proteins in determining the fast reactivity and rate order of Trx. A reaction mechanism involving the transient formation of a Trx-protein binding complex is proposed for the oxidoreduction of protein thiols-disulfides. Furthermore, studies on insulin reduction show that Trx acts as an enzyme rather than a redox couple. These results provide explanations for the observed variations of the CXXC motif in PDI-like proteins as well as the conservation of the CXXC motif in Trx.
Simple sequence repeat variations expedite phage divergence: Mechanisms of indels and gene mutations
(2016-07)
Lin, Tiao-Yin
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生物科技學系
;
Department of Biological Science and Technology
Phages are the most abundant biological entities and influence prokaryotic communities on Earth. Comparing closely related genomes sheds light on molecular events shaping phage evolution. Simple sequence repeat (SSR) variations impart over half of the genomic changes between T7M and T3, indicating an important role of SSRs in accelerating phage genetic divergence. Differences in coding and noncoding regions of phages infecting different hosts, coliphages T7M and T3, Yersinia phage phi YeO3-12, and Salmonella phage phi SG-JL2, frequently arise from SSR variations. Such variations modify noncoding and coding regions; the latter efficiently changes multiple amino acids, thereby hastening protein evolution. Four classes of events are found to drive SSR variations: insertion/deletion of SSR units, expansion/contraction of SSRs without alteration of genome length, changes of repeat motifs, and generation/loss of repeats. The categorization demonstrates the ways SSRs mutate in genomes during phage evolution. Indels are common constituents of genome variations and human diseases, yet, how they occur without preexisting repeat sequence is less understood. Non-repeat-unit-based misalignment-elongation (NRUBME) is proposed to be one mechanism for indels without adjacent repeats. NRUBME or consecutive NRUBME may also change repeat motifs or generate new repeats. NRUBME invoking a non-Watson-Crick base pair explains insertions that initiate mononucleotide repeats. Furthermore, NRUBME successfully interprets many inexplicable human di- to tetranucleotide repeat generations. This study provides the first evidence of SSR variations expediting phage divergence, and enables insights into the events and mechanisms of genome evolution. NRUBME allows us to emulate natural evolution to design indels for various applications. (C) 2016 Elsevier B.V. All rights reserved.
Truncated Escherichia coli thioredoxin induces proliferation of human blood mononuclear cells and production of reactive oxygen species as well as proinflammatory cytokines
(2016-06)
Liu, Si-Yen
;
Liu, I-Chung
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Lin, Tiao-Yin
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生物科技學系
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分子醫學與生物工程研究所
Thioredoxin (Trx) is a redox protein characterized by a Trx fold. A naturally occurring truncated human Trx, Trx 80, which lacks the C-terminal strand-helix of the Trx fold, stimulates proliferation of peripheral blood mononuclear cells (PBMCs). It has not been clear how Trx80 gains this function. This study investigates whether a peptide with substantial sequence difference from Trx80, but retaining an abridged Trx fold can elicit PBMC proliferation. We genetically truncated a carboxy-terminal beta-alpha motif of Escherichia coli Trx to produce a peptide, Trx83, which shares low sequence identity with human Trx80. Addition of reduced-form Trx83 to resting human PBMCs promoted cell proliferation, while oxidized-form Trx83 lacked the function. By contrast, oxidized-form Trx80 exhibited a high activity in promoting PBMC proliferation, indicating the importance of sequence context of an abridged thioredoxin in influencing PBMC proliferation. Trx83 increases cellular reactive oxygen species and proinflammatory cytokines TNF-alpha and IL-1 beta, suggesting that Trx83 modulates inflammatory pathways. This notion is supported by the observation that cystine or cysteine abolishes the Trx83 induced PBMC proliferation. The PBMC stimulatory activity of Trx83 may have potential for pharmacological developments. SIGNIFICANCE OF THE STUDY Elicitation of primary proliferative responses of PBMCs by a protein is generally difficult. We show that Escherichia coli Trx83 with a truncated Trx fold induces PBMC proliferation, but only in the disulfide-reduced form. In contrast, oxidized-form human Trx80 is a potent stimulator. These results demonstrate that the sequence context of an abridged Trx fold is influential in inducing PBMC proliferation. The stimulatory effect of Trx83 is associated with an increase of inflammatory response. The possibility of eliciting PBMC proliferation and switching this activity on/off by redox control provides a perspective for developing Trx83 as a PBMC stimulatory agent. Copyright (C) 2016 John Wiley & Sons, Ltd.