Comparison of the Structure and Biological Activities of Wild-type and Mutant Liver-targeting Peptide Modified Recombinant Human Endostatin (rES-CSP) in Human Hepatocellular Carcinoma HepG2 Cells.
A missense mutation of Aspartic to Asparagine acid in 113 position of liver-targeting peptide CSP I-plus modified rEndostatin (rES-CSP) happened unexpectedly results in the changes of protein secondary structure and a reduced bioactivity. With the aim to clarify the structure-function relationships featuring the fuse protein rES-CSP, the three-dimensional structural models of wild-type and mutant D113NrES-CSP were constructed by template-based modeling approach. To evaluate the effect of the single mutation on rES-CSP stability, the molecular dynamic simulation was used to reveal the structural and dynamic characteristics. Analysis on the bioactivity were conducted using a number of validated in vitro assays including proliferation, migration, cell cycle and apoptosis in HepG2 cells. Results showed that the mutant rES-CSP reduce the stability and loss of function, and the wild-type rES-CSP could both bind to the normal liver cells Chang's and the hepatoma cells HepG2 but significantly higher than non-targeted rEndostatin. rESCSP could inhibit the proliferation of hepatoma cells in a dose-dependent manner, and increase the proportion of G1 phase, reduce the proportion of S phase, promote the apoptosis on hepatoma cells. These results make a further complement of the mechanisms by which the fuse protein rES-CSP would provide a feasible and convenient approach to produce liver-targeting drugs for treatment of the liver disease.
Dongmei
D
Bao
Xiaobao
X
Jin
Yan
Y
Ma
Guangdong Provincial Key Laboratory of Pharmaceutical Bioactive Substances, Guangdong Pharmaceutical University, Guangzhou, People's Republic of China. zhujy@gdpu.edu.cn.
Jiayong
J
Zhu
D000595:Amino Acid Sequence; D017209:Apoptosis / Q000187:drug effects; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D002453:Cell Cycle / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D043169:Endostatins / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology*; D056945:Hep G2 Cells; D006801:Humans; D008099:Liver / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D008113:Liver Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D010455:Peptides / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology*; D011487:Protein Conformation; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology
D043169:Endostatins; D010455:Peptides; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; C041595:circumsporozoite protein, Protozoan