ErbB3和ErbB4特异性配体筛选系统的建立及neuregulin-1突变体的筛选

doi:10.3724/SP.J.1005.2010.01247

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HEREDITAS ›› 2010, Vol. 32 ›› Issue (12): 1247-1255.doi: 10.3724/SP.J.1005.2010.01247

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Establishment of an ErbB3- and ErbB4-specific ligand screening system, and a screen of neuregulin-1 mutants

QI Ying^{1, 2}, WEI Dong-Zhi¹, LIU Xi-Fu², ZHOU Ming-Dong²

1. Newworld Institute of Biotechnology, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China;
2. Zensun (Shanghai) Science and Technology, Ltd., Shanghai 201203, China

Received:2010-03-29 Revised:2010-06-12 Online:2010-12-20 Published:2010-12-20
Contact: WEI Dong-Zhi;ZHOU Ming-Dong2 E-mail:dzhwei@ecust.edu.cn；mdzhou@zensun.com

Abstract

Abstract: Neuregulin-1 (NRG1), now in a phase II clinical trial, has beneficial effects on heart failure patients through the activation of ErbB2/ErbB4 receptor pair. To decrease the side effect of NRG1 on activating ErbB3, a mutation screen was carried out to get NRG1 mutants, which have more specific binding to ErbB2/ErbB4 receptor pair. Two CHO stable cell lines were constructed, which express ErbB2/ErbB3 or ErbB2/ErbB4 receptor pair, respectively. The ErbB2/ErbB4 cell line showed similar characteristics in ligand-binding activity and the activation of downstream signaling molecules, such as the AKT and PI3K to the primary neonatal rat ventricular myocytes (NRVM), which endogenously expresses ErbB2/ErbB4. Both cell lines have good dose-response. Thirty-one NRG1 mutants were successfully expressed in Escherichia coli and purified. Their ability to stimulate the downstream signaling was measured by detecting AKT phosphorylation. Some mu-tants showed more specific activation activity in ErbB2/ErbB4 cells. Further study on five of these mutants demonstrated that the change of the activation activity is associated with that of their binding activities to ErbB2/ErbB4 and ErbB2/ErbB3. Four of the candidates are more specific ligands for ErbB2/ErbB4 activation, and thus may serve as more potent drug can-didates for heart failure.

Key words: NRG1 mutant, heart failure, stable transfected cell line, ErbB family, p-AKT

CU Yang, WEI Dong-Zhi, LIU Chi-Fu, ZHOU Meng-Dong. Establishment of an ErbB3- and ErbB4-specific ligand screening system, and a screen of neuregulin-1 mutants[J]. HEREDITAS, 2010, 32(12): 1247-1255.

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Establishment of an ErbB3- and ErbB4-specific ligand screening system, and a screen of neuregulin-1 mutants

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