Synthesis of functional proteins using Escherichia coli extract entrapped in calcium alginate microbeads.
In this report, we describe the construction and analysis of a cell-free protein synthesis system immobilized in calcium alginate microbeads. When incubated in a feeding solution that contained amino acids and other low-molecular-weight substrates, the microbeads transcribed and translated coimmobilized DNA into functional proteins. Protein synthesis continued for more than 15 h with the diffusional supply of substrates and removal of by-products. In addition, functional proteins were generated from PCR-amplified genes as efficiently as from plasmid, suggesting that these cell-like microbeads could be used for functional screening of genomic libraries.
Department of Fine Chemical Engineering and Chemistry, Chungnam National University, Daejeon 305-764, Korea.
Yong-Chan
YC
Kwon
Geun-Hee
GH
Hahn
Kang Moo
KM
Huh
Dong-Myung
DM
Kim
D000464:Alginates / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000096:biosynthesis*; D020723:Glucuronic Acid / Q000737:chemistry; D049452:Green Fluorescent Proteins / Q000096:biosynthesis; D006603:Hexuronic Acids / Q000737:chemistry; D008863:Microspheres
D000464:Alginates; D029968:Escherichia coli Proteins; D006603:Hexuronic Acids; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D020723:Glucuronic Acid