Engineering a pharmacologically superior form of granulocyte-colony-stimulating factor by fusion with gelatin-like-protein polymer.
The plasma half-life of therapeutic proteins is a critical factor in many clinical applications. Therefore, new strategies to prolong plasma half-life of long-acting peptides and protein drugs are in high demand. Here, we designed an artificial gelatin-like protein (GLK) and fused this hydrophilic GLK polymer to granulocyte-colony-stimulating factor (G-CSF) to generate a chimeric GLK/G-CSF fusion protein. The genetically engineered recombinant GLK/G-CSF (rGLK/G-CSF) fusion protein was purified from Pichia pastoris. In vitro studies demonstrated that rGLK/G-CSF possessed an enlarged hydrodynamic radius, improved thermal stability and retained full bioactivity compared to unfused G-CSF. Following a single subcutaneous administration to rats, the rGLK/G-CSF fusion protein displayed a slower plasma clearance rate and stimulated greater and longer lasting increases in circulating white blood cells than G-CSF. Our findings indicate that fusion with this artificial, hydrophilic, GLK polymer provides many advantages in the construction of a potent hematopoietic factor with extended plasma half-life. This approach could be easily applied to other therapeutic proteins and have important clinical applications.
Huang
Yan-Shan
YS
Department of Cell Biology, Zhejiang University, Hangzhou, China.
Wen
Xiao-Fang
XF
Wu
Yi-Liang
YL
Wang
Ye-Fei
YF
Fan
Min
M
Yang
Zhi-Yu
ZY
Liu
Wei
W
Zhou
Lin-Fu
LF
D000595:Amino Acid Sequence; D000818:Animals; D000917:Antibody Formation / Q000187:drug effects / Q000276:immunology; D001483:Base Sequence; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004355:Drug Stability; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005780:Gelatin / Q000737:chemistry*; D016179:Granulocyte Colony-Stimulating Factor / Q000009:adverse effects / Q000097:blood / Q000737:chemistry / Q000235:genetics* / Q000494:pharmacology; D006207:Half-Life; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010843:Pichia / Q000235:genetics; D011108:Polymers / Q000737:chemistry*; D015202:Protein Engineering; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000009:adverse effects / Q000097:blood / Q000737:chemistry / Q000235:genetics* / Q000494:pharmacology; D011994:Recombinant Proteins; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014162:Transfection
D011108:Polymers; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D016179:Granulocyte Colony-Stimulating Factor; D005780:Gelatin