Cloning and heterologous expression of a hydrophobin gene Ltr.hyd from the tiger milk mushroom Lentinus tuber-regium in yeast-like cells of Tremella fuciformis
Background: Hydrophobins are small proteins secreted by filamentous fungi, which show a highly surface activity. Because of the signally self-assembling abilities and surface activities, hydrophobins were considered as candidates in many aspects, for example, stabilizing foams and emulsions in food...
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doaj-3ebb4a8081ce4c75bb98cb1640b099012020-11-25T01:46:05ZengElsevierElectronic Journal of Biotechnology0717-34582018-03-0132612Cloning and heterologous expression of a hydrophobin gene Ltr.hyd from the tiger milk mushroom Lentinus tuber-regium in yeast-like cells of Tremella fuciformisDongmei Liu0Hanyu Zhu1Yue Chen2Liesheng Zheng3Liguo Chen4Aimin Ma5College of Food Science and Technology, Huazhong Agricultural University, Wuhan 430070, ChinaCollege of Food Science and Technology, Huazhong Agricultural University, Wuhan 430070, ChinaCollege of Food Science and Technology, Huazhong Agricultural University, Wuhan 430070, ChinaCollege of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070, ChinaCollege of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070, ChinaCollege of Food Science and Technology, Huazhong Agricultural University, Wuhan 430070, China; Key Laboratory of Agro-Microbial Resources and Utilization, Ministry of Agriculture, Huazhong Agricultural University, Wuhan 430070, China; Corresponding author.Background: Hydrophobins are small proteins secreted by filamentous fungi, which show a highly surface activity. Because of the signally self-assembling abilities and surface activities, hydrophobins were considered as candidates in many aspects, for example, stabilizing foams and emulsions in food products. Lentinus tuber-regium, known as tiger milk mushroom, is both an edible and medicinal sclerotium-producing mushroom. Up to now, the hydrophobins of L. tuber-regium have not been identified. Results: In this paper, a Class I hydrophobin gene, Ltr.hyd, was cloned from L. tuber-regium and expressed in the yeast-like cells of Tremella fuciformis mediated by Agrobacterium tumefaciens. The expression vector pGEH-GH was under the control of T. fuciformis glyceraldehyde-3-phosphate dehydrogenase gene (gpd) promoter. The integration of Ltr.hyd into the genome of T. fuciformis was confirmed by PCR, Southern blot, fluorescence observation and quantitative real-time PCR (qRT-PCR). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that recombinant hydrophobin rLtr.HYD with an expected molecular mass of 13 kDa was extracted. The yield of rLtr.HYD was 0.66 mg/g dry weight. The emulsifying activity of rLtr.HYD was better than the typical food emulsifiers sodium caseinate and Tween 20. Conclusions: We evaluated the emulsifying property of hydrophobin Ltr.HYD, which can be potentially used as a food emulsifier. Keywords: Agrobacterium tumefaciens, Emulsifier, Expression vector, Filamentous fungi, Gel electrophoresis, Glyceraldehyde-3-phosphate dehydrogenase, Heterogenous expression, Hydrophobin, Quantitative real-time PCR, Southern blot, Surface activityhttp://www.sciencedirect.com/science/article/pii/S0717345817300817 |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Dongmei Liu Hanyu Zhu Yue Chen Liesheng Zheng Liguo Chen Aimin Ma |
spellingShingle |
Dongmei Liu Hanyu Zhu Yue Chen Liesheng Zheng Liguo Chen Aimin Ma Cloning and heterologous expression of a hydrophobin gene Ltr.hyd from the tiger milk mushroom Lentinus tuber-regium in yeast-like cells of Tremella fuciformis Electronic Journal of Biotechnology |
author_facet |
Dongmei Liu Hanyu Zhu Yue Chen Liesheng Zheng Liguo Chen Aimin Ma |
author_sort |
Dongmei Liu |
title |
Cloning and heterologous expression of a hydrophobin gene Ltr.hyd from the tiger milk mushroom Lentinus tuber-regium in yeast-like cells of Tremella fuciformis |
title_short |
Cloning and heterologous expression of a hydrophobin gene Ltr.hyd from the tiger milk mushroom Lentinus tuber-regium in yeast-like cells of Tremella fuciformis |
title_full |
Cloning and heterologous expression of a hydrophobin gene Ltr.hyd from the tiger milk mushroom Lentinus tuber-regium in yeast-like cells of Tremella fuciformis |
title_fullStr |
Cloning and heterologous expression of a hydrophobin gene Ltr.hyd from the tiger milk mushroom Lentinus tuber-regium in yeast-like cells of Tremella fuciformis |
title_full_unstemmed |
Cloning and heterologous expression of a hydrophobin gene Ltr.hyd from the tiger milk mushroom Lentinus tuber-regium in yeast-like cells of Tremella fuciformis |
title_sort |
cloning and heterologous expression of a hydrophobin gene ltr.hyd from the tiger milk mushroom lentinus tuber-regium in yeast-like cells of tremella fuciformis |
publisher |
Elsevier |
series |
Electronic Journal of Biotechnology |
issn |
0717-3458 |
publishDate |
2018-03-01 |
description |
Background: Hydrophobins are small proteins secreted by filamentous fungi, which show a highly surface activity. Because of the signally self-assembling abilities and surface activities, hydrophobins were considered as candidates in many aspects, for example, stabilizing foams and emulsions in food products. Lentinus tuber-regium, known as tiger milk mushroom, is both an edible and medicinal sclerotium-producing mushroom. Up to now, the hydrophobins of L. tuber-regium have not been identified. Results: In this paper, a Class I hydrophobin gene, Ltr.hyd, was cloned from L. tuber-regium and expressed in the yeast-like cells of Tremella fuciformis mediated by Agrobacterium tumefaciens. The expression vector pGEH-GH was under the control of T. fuciformis glyceraldehyde-3-phosphate dehydrogenase gene (gpd) promoter. The integration of Ltr.hyd into the genome of T. fuciformis was confirmed by PCR, Southern blot, fluorescence observation and quantitative real-time PCR (qRT-PCR). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that recombinant hydrophobin rLtr.HYD with an expected molecular mass of 13 kDa was extracted. The yield of rLtr.HYD was 0.66 mg/g dry weight. The emulsifying activity of rLtr.HYD was better than the typical food emulsifiers sodium caseinate and Tween 20. Conclusions: We evaluated the emulsifying property of hydrophobin Ltr.HYD, which can be potentially used as a food emulsifier. Keywords: Agrobacterium tumefaciens, Emulsifier, Expression vector, Filamentous fungi, Gel electrophoresis, Glyceraldehyde-3-phosphate dehydrogenase, Heterogenous expression, Hydrophobin, Quantitative real-time PCR, Southern blot, Surface activity |
url |
http://www.sciencedirect.com/science/article/pii/S0717345817300817 |
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