Cloning, Expression, Characterization, and Tissue Distribution of Cystatin C from Silver Carp (Hypophthalmichthys molitrix)

Li Ran, Xiaoqian Tan, Shuhong Li, Yu Jin, Song Li, Shulei Li, Timo Takala, Per Saris

Research output: Contribution to journalArticleScientificpeer-review

Abstract

Cystatins are proteins, which inhibit cysteine proteases, such as papain. In this study, the 336-bp cystatin C gene (family II, HmCysC) of silver carp (Hypophthalmichthys molitrix) was cloned and expressed in Escherichia coli BL21 (DE3). HmCysC encodes the mature peptide of cystatin C (HmCystatin C), with 111 amino acids. A typical QXXXG motif was found in HmCystatin C and it formed a cluster with Cyprinus carpio and Danio rerio cystatin C in the phylogenetic tree. Quantitative real-time polymerase chain reaction analysis indicated that HmCysC was transcribed at different levels in five tested tissues of silver carp. Following purification with Ni2+– nitrilotriacetic acid agarose affinity chromatography, HmCystatin C displayed a molecular weight of 20 kDa in sodium dodecyl sulfate polyacrylamide gel electrophoresis. Purified HmCystatin C had strong inhibitory effects toward the proteolytic activity of papain. Immunochemical staining with anti-HmCystatin C antibody showed that HmCystatin C was widely distributed in silver carp tissues. These results collectively demonstrated the properties of HmCystatin C, providing information for further studies of cystatins from fish organisms.
Original languageEnglish
JournalJournal of Agricultural and Food Chemistry
Volume69
Issue number17
Pages (from-to)5144–5154
Number of pages11
ISSN0021-8561
DOIs
Publication statusPublished - 21 Apr 2021
MoE publication typeA1 Journal article-refereed

Fields of Science

  • 416 Food Science
  • 1182 Biochemistry, cell and molecular biology
  • silver carp (Hypophthalmichthys molitrix)
  • cystatin C
  • cloning
  • transcription level
  • tissue distribution
  • protein quantification
  • CYSTEINE PROTEINASE-INHIBITOR
  • AMINO-ACID-SEQUENCE
  • MOLECULAR-CLONING
  • CATHEPSIN-B
  • OVARIAN CYSTATIN
  • GENE-EXPRESSION
  • EGG-WHITE
  • PURIFICATION
  • IDENTIFICATION
  • MUSCLE

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