Analytical Methods
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2019
·Vol. 11
·期 16
·2197-2203
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IF 2.378
Xiaoqiang Wang,Haiyan Sun,Chengkun Liu,Chao Wang
Following an inspiration from the chemistry of protein nanocages in nature, we evaluated the performance of a hemin-functionalized GroEL nanocage as peroxidase mimics for chromogenic detection. The reactivity of this biohybrid formed by detergent dialysis was found to be highly dependent on the amount of hemin encapsulated inside the GroEL central cavity. In comparison with horseradish peroxidase (HRP), the hemin–GroEL complex exhibited a higher recognition ability and reactivity towards the chromogenic substrate tested, while, similar to HRP, the peroxidase mimic followed a ping-pong catalytic mechanism. Moreover, the efficacy of hemin–GroEL for the chromogenic detection of H2O2 or glucose was demonstrated, with the detection limits determined to be 7 μM for H2O2 and 11 μM for glucose. This assay was also shown to be useful for the oxidative removal of phenolic compounds. Thus, the GroEL cage is a valuable tool for the development of analytical nano-materials and practical applications such as clinical analysis and environmental monitoring.
研究领域:
生物化学
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纳米技术
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分析化学
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环境科学
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DOI
高影响因子
BIOTECHNOLOGY LETTERS
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2023
·Vol. 45
·期 8
·993-1000
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IF 2.7
Peng Shuaiying,Chu Zhongmei,Lu Jianfeng,Li Dongxiao,Wang Yonghong,Yang Shengli,Zhang Yi
The self-sufficient cytochrome P450 BM3 mutant (A74G/F87V/D168H/L188Q) can serve as a biocatalyst for whole-cell catalysis process of indigo. Nevertheless, the bioconversion yield of indigo is generally low under normal cultivation conditions (37 °C, 250 rpm). In this study, a recombinant E. coli BL21(DE3) strain was constructed to co-express the P450 BM3 mutant gene and GroEL/ES genes to investigate whether GroEL/ES can promote the indigo bioconversion yield in E. coli . The results revealed that the GroEL/ES system could significantly increase the indigo bioconversion yield, and the indigo bioconversion yield of the strain co-expressing P450 BM3 mutant and GroEL/ES was about 21-fold that of the strain only expressing the P450 BM3 mutant. In addition, the P450 BM3 enzyme content and in vitro indigo bioconversion yield were determined to explore the underlying mechanism for the improvement of indigo bioconversion yield. The results revealed that GroEL/ES did not increase indigo bioconversion yield by increasing the content of P450 BM3 enzyme and its enzymatic transformation efficiency. Moreover, GroEL/ES could improve the intracellular nicotinamide adenine dinucleotide phosphate (NADPH)/NADP + ratio. Given that NADPH is an important coenzyme in the catalytic process of indigo, the underlying mechanism for the improvement of indigo bioconversion yield is probably related to an increase in the intracellular NADPH/NADP + ratio.
研究领域:
生物化学
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生物技术
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酶学
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DOI
PubMed
高影响因子
ACS Applied Materials & Interfaces
·
2017
·
IF 7.504
Xiaoqiang Wang,Chao Wang,Meihong Pan,Junting Wei,Fuping Jiang,Rongsheng Lu,Xuan Liu,Yihui Huang,Fang Huang
Taking inspiration from biology’s effectiveness in functionalizing protein-based nanocages for chemical processes, we describe here a rational design of an artificial metalloenzyme for oxidations with the bacterial chaperonin GroEL, a nanocage for protein folding in nature, by supramolecular anchoring of catalytically active hemin in its hydrophobic central cavity. The promiscuity of the chaperonin cavity is an essential element of this design, which can mimic the hydrophobic binding pocket in natural metalloenzymes to accept cofactor and substrate without requiring specific ligand–protein interactions. The success of this approach is manifested in the efficient loading of multiple monomeric hemin cofactors to the GroEL cavity by detergent dialysis and good catalytic oxidation properties of the resulting biohybrid in tandem with those of the clean oxidant of H2O2. Investigation of the mechanism of hemin–GroEL-catalyzed oxidation of two-model substrates reveals that the kinetic behavior of the complex follows a ping-pong mechanism in both cases. Through comparison with horseradish peroxidase, the oxidative activity and stability of hemin–GroEL were observed to be similar to those found in natural peroxidases. Adenosine 5′-triphosphate (ATP)-regulated partial dissociation of the biohybrid, as assessed by the reduction of its catalytic activity with the addition of the nucleotide, raises the prospect that ATP may be used to recycle the chaperonin scaffold. Moreover, hemin–GroEL can be applied to the chromogenic detection of H2O2, which (or peroxide in general) is commonly contained in industrial wastes. Considering the rich chemistry of free metalloporphyrins and the ease of production of GroEL and its supramolecular complex with hemin, this work should seed the creation of many new artificial metalloenzymes with diverse reactivities.
研究领域:
生物化学
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酶工程
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催化化学
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DOI
PubMed
高影响因子
ANALYTICAL AND BIOANALYTICAL CHEMISTRY
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2019
·Vol. 411
·期 17
·3819-3827
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IF 3.286
Wang Xiaoqiang,Sun Haiyan,Wang Chao
The design of functionalized protein nanocages as enzyme mimics is a relatively new and promising field as these biogenic nanocapsules are inherently monodisperse with precise architectures, unattainable by purely synthetic processes. This work explored the catalytic properties of the bacterial GroEL-nanocaged hemin and its performance in label-free colorimetric assays. It is demonstrated that the hemin-GroEL biohybrid has peroxidase-like activity and follows the typical Michaelis-Menten kinetics and ping-pong mechanism in the model sensing processes. The open nature and nanoreactor effect of the GroEL cage and the addition of ATP are shown to significantly influence the catalytic activity. For glucose detection with the hemin-GroEL complex, the linearity between the analyte concentration and UV-vis absorption was determined to range from 0 to 200 μM with a limit of detection (LOD) of ~ 12 μM under the defined conditions. In addition, the colorimetric detection of catechol led to a linear dynamic range of 0–120 μM with a LOD of ~ 17 μM. This indicates that the same assay could be used as a sensing platform for the detection or even oxidative removal of phenolic contaminants in the presence of H 2 O 2 . Thus, GroEL cage is a valuable tool for the development of nanozymes and practical applications such as clinical analysis and environmental monitoring.
研究领域:
生物化学
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纳米技术
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分析化学
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生物技术
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DOI
PubMed
高影响因子
SPECTROCHIMICA ACTA PART A-MOLECULAR AND BIOMOLECULAR SPECTROSCOPY
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2020
·
IF 3.232
Xiaoqiang Wang,Zhuang Shi,Han Chen,Fang Huang
In this paper, we introduce a new type of functional, supramolecular porphyrin conjugate created using the bacterial GroEL protein cage based on non-specific hydrophobic interaction. The synthesis, structure and property of the porphyrin conjugate were characterized by dynamic light scattering , UV–vis spectroscopy and fluorescence spectroscopy. We observed that the model zinc-tetraphenylporphyrin (Zn-TPP) with high hydrophobicity can be well-dispersed in aqueous solutions with the aid of GroEL open chamber, which is known to be a favorable nanocompartment for aggregation-prone molecules. The maximal encapsulation efficiency of Zn-TPP in GroEL was determined to be ~98%. It is further seen that the constructed double Zn-TPP-GroEL complex exhibited good photocatalytic activity in the model reactions of the production of singlet oxygen and the reduction of methyl viologen under illumination with visible light. Moreover, we found that GroEL can significantly improve the photostability of Zn-TPP molecules as a result of nanoscale assembly within its hydrophobic chamber. Hence enhanced water solubility and photostability of Zn-TPP, which are considered as the first two hurdles for the wide usage of porphyrins, were achieved simultaneously by the development of GroEL cage as a building block. Supramolecular nanostructures formed from porphyrins (or related molecules) and GroEL for photocatalysis would greatly simplify applications of such structures.
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PubMed
高影响因子
PROCESS BIOCHEMISTRY
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2017
·
IF 2.497
Hui Zhang,Mengya Li,Jie Li,Guangli Wang,Feng Li,Minghua Xiong
Nitrile hydratase (NHase) has attracted substantial attention for industrial applications to produce large-scale amides. Several NHases have been investigated for functional expression in Escherichia coli ( E. coli ). A Fe-type NHase was obtained from an acetamiprid-degrading bacterium, Pseudoxanthomonas sp. AAP-7 and functionally expressed in E. coli BL21 (DE3). No significant NHase activity was detected from the E. coli expressing either the NHase gene alone or NHase and P46K genes transcribed as one unit. Purified recombinant NHase, co-expressed with P46K on two separate plasmids, exhibited the maximal enzyme activity. Furthermore, a GST tag attached to the N-terminus of α subunit resulted in a slight increase in the solubility and stability of NHase compared with a His tag at the C-terminus of β subunit. When co-expressed with the chaperones GroEL-GroES, the yield of the soluble recombinant NHase was improved substantially, while a small decrease in NHase activity was observed. The putative activator P46K was strictly required for production of the recombinant NHase for full enzyme activity, although the chaperones GroEL-GroES appeared to assist NHase to fold properly. This study of the expression of a fully active Fe-type NHase would provide another example to enhance our understanding of NHase biosynthesis.
研究领域:
生物化学
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微生物学
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酶学
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工业生物技术
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DOI
高影响因子
JOURNAL OF MATERIALS SCIENCE
·
2018
·Vol. 53
·期 12
·8786-8794
·
IF 2.993
Wang Xiaoqiang,Xu Baomei,Liu Zhenzhen
Supramolecular anchoring of metalloporphyrins in a protein is an attractive approach to the generation of artificial enzymes. Here, we employ the hydrophobic nanocage of single-ring mutant of bacterial GroEL protein for this purpose. We found that multiple monomeric hemin cofactors can be efficiently loaded into the protein nanocage. The as-prepared biohybrid possessed an oxidase-like catalytic activity and followed the typical Michaelis–Menten kinetics and a ping-pong mechanism in the H 2 O 2 -mediated oxidation of model substrates. In comparison with natural peroxidase, the artificial enzyme exhibited higher affinity for the model substrate. A simple and sensitive colorimetric method for the quantitative detection of H 2 O 2 and glucose was also developed based on the artificial enzyme, with the detection limits determined to be 3.0 μM for H 2 O 2 and 5.0 μM for glucose, respectively. The protein nanocage-based artificial enzyme is very flexible and is envisioned to be adapted readily for binding other metal complexes and catalysis of other reactions.
研究领域:
生物化学
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酶学
·
分析化学
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DOI
高影响因子
MATERIALS & DESIGN
·
2020
·
IF 6.289
Xiaoqiang Wang,Chengkun Liu,Zhuang Shi,Meihong Pan,Daoyong Yu
This paper reports a novel design of an artificial pigment-protein complex, created by supramolecular binding of chlorophyll a (Chl a ) in the hydrophobic cavity of s ingle- r ing mutant of bacterial GroEL (SR). The results show that Chl a molecules assembled in SR nano-ring retain their native structure, and an increase of loading amount (up to ~26 pigment molecules per protein nano-ring) with the equilibrium concentration of Chl a upon dialysis has been observed. It is further seen that the nanoscale organization of Chl a in SR leads to a significant quenching of Chl a fluorescence, especially at high loadings. Moreover, the photostability of Chl a encapsulated in SR is significantly enhanced in comparison with free Chl a , as evidenced by an up to 6-fold increase of its half-life time. Both the interaction of Chl a with SR and the interaction between Chl a molecules contribute to photostability. Finally, it is shown that the Chl a -SR complex has good photoreduction activity as well as photoelectrochemical activity under visible light illumination. The maximum photoelectric conversion efficiency of the Chl a -SR-sensitized solar cells is around 1.09%. The strategy presented herein is believed to be instrumental in the design of effective photoenergy conversion materials.
研究领域:
生物物理学
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生物化学
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材料科学
·
可再生能源
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产品关联
DOI
高影响因子
危险品化学品经营许可证（带存储）