Expression, immobilization and enzymatic properties of glutamate decarboxylase fused to a cellulose-binding domain.
Escherichia coli-derived glutamate decarboxylase (GAD), an enzyme that catalyzes the conversion of glutamic acid to gamma-aminobutyric acid (GABA), was fused to the cellulose-binding domain (CBD) and a linker of Trichoderma harzianum endoglucanase II. To prevent proteolysis of the fusion protein, the native linker was replaced with a S(3)N(10) peptide known to be completely resistant to E. coli endopeptidase. The CBD-GAD expressed in E. coli was successfully immobilized on Avicel, a crystalline cellulose, with binding capacity of 33 ± 2 nmol(CBD-GAD)/g(Avicel) and the immobilized enzymes retained 60% of their initial activities after 10 uses. The results of this report provide a feasible alternative to produce GABA using immobilized GAD through fusion to CBD.
Biotechnology Process Engineering Center, KRIBB, Daejeon 305-600, Korea; E-Mails:  gohworld@kribb.re.kr  (H.P.);  ahnjo@kribb.re.kr  (J.A.);  jhlee@kribb.re.kr  (J.L.);  tntn7616@kribb.re.kr  (H.L.);  chskim@kribb.re.kr  (C.K.);  jkjung@kribb.re.kr  (J.-K.J.);  hwlee@kribb.re.kr  (H.L.).
Hyemin
H
Park
Jungoh
J
Ahn
Juwhan
J
Lee
Hyeokwon
H
Lee
Chunsuk
C
Kim
Joon-Ki
JK
Jung
Hongweon
H
Lee
Eun Gyo
EG
Lee
D002480:Cellulase / Q000737:chemistry / Q000378:metabolism; D002482:Cellulose / Q000737:chemistry / Q000378:metabolism*; D010450:Endopeptidases / Q000378:metabolism; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000378:metabolism; D005968:Glutamate Decarboxylase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D059748:Proteolysis; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D014242:Trichoderma / Q000201:enzymology; D005680:gamma-Aminobutyric Acid / Q000378:metabolism
D004800:Enzymes, Immobilized; D010455:Peptides; D011993:Recombinant Fusion Proteins; D005680:gamma-Aminobutyric Acid; D002482:Cellulose; C056373:endoglucanase 2; D002480:Cellulase; D010450:Endopeptidases; D005968:Glutamate Decarboxylase