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A Novel Targeted Multi-Functional Fusion Protein Possesses Inhibitory Activities Against Bacteria, Thrombin and Platelet Aggregation

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Abstract

In the present study, we designed a novel targeted multi-functional fusion protein LHAD composed of LL-37, FXa recognition peptide, hirudin-12-residue, AAP, and RGD peptide. It was expressed in the Pichia pastoris GS115 strain and purified by affinity chromatography. The in vitro studies suggested that the novel designed protein exhibited antibacterial activity, anti-platelet aggregation and anti-thrombin activities. Moreover, the capability of anti-thrombin was significantly increased compared to that of natural hirudin. Our study may provide a potential approach to design multi-functional drugs for the prevention and management of thrombosis.

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Abbreviations

VEC:

Vascular endothelial cell

RGD:

Arg-Gly-Asp

AAP:

pGlu-Asn-Trp

EK:

Enterokinase

FXa:

Activated factor X

Amp:

Ampicillin

ADP:

Adenosine diphosphate

ATU:

Antithrombin unit

kDa:

Kilo-dalton

PT:

Prothrombin time

APTT:

Activated partial thromboplastin time

GpIIb/IIIa:

Glycol-protein IIb/IIIa

PBS:

Phosphate buffered saline

PBST:

Phosphate buffered saline Tween-20

LDAO:

Lauryldimethylamine N-oxide

Ni-IDA:

Nickel-iminidoacetic acid

BSA:

Bovine serum albumin

DAB:

Diaminobenzidine

SPSS:

Statistical product and service solutions

BMM:

Buffered methanol medium

BMMY:

Buffered methanol-complex medium

LB:

Luria Bertani medium

PRP:

Platelet rich plasma

PPP:

Platelet-poor plasma

SOE PCR:

Splicing by overlap extension PCR

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Acknowledgments

This work was supported by key scientific and technological project of Chongqing (NO: CSTC, 2009AB5174), and by the Fundamental Research Funds for the Central Universities (NO: CDJZR10230018). We thank Prof. Donald Grierson (Nottingham University) for critical reading of the manuscript.

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Authors and Affiliations

  1. Bioengineering College, Chongqing University, Chongqing, 400044, People’s Republic of China

    Jiange Gao, Zongli Hu, Zhiping Zhao, Guanglei Liu & Guoping Chen

  2. Department of Biological and Chemical Engineering, Chongqing Education College, Chongqing, 400067, People’s Republic of China

    Yanrong Ren

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  1. Jiange Gao
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  2. Zongli Hu
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  3. Zhiping Zhao
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  4. Guanglei Liu
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  5. Yanrong Ren
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Correspondence to Zongli Hu.

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Gao, J., Hu, Z., Zhao, Z. et al. A Novel Targeted Multi-Functional Fusion Protein Possesses Inhibitory Activities Against Bacteria, Thrombin and Platelet Aggregation. Protein J 30, 521–528 (2011). https://doi.org/10.1007/s10930-011-9357-6

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