|
|
Molecular Biology and Genetics |
| Electronic Journal of Biotechnology ISSN: 0717-3458 |
Vol.
13 No. 3, Issue of May 15, 2010 |
| © 2010 by Pontificia Universidad Católica
de Valparaíso -- Chile |
Received April 6, 2009
/ Accepted December 15, 2009 |
| DOI: 10.2225/vol13-issue3-fulltext-8 |
|
Human
sulfatase transiently and functionally active expressed in E. coli K12
Raúl A. Poutou-Piñales
Instituto de Errores Innatos
del Metabolismo
Grupo de Biotecnología
Ambiental e Industrial
Facultad de Ciencias
Pontificia Universidad
Javeriana
Cra. 7 N° 40-62. Bogotá.
D.C., Colombia
E-mail: rpoutou@javeriana.edu.co
Adriana Vanegas Niño
Instituto de Errores Innatos
del Metabolismo
Facultad de Ciencias
Pontificia Universidad
Javeriana
Cra. 7 N° 40-62. Bogotá.
D.C., Colombia
Patricia Landázuri
Laboratorio de Bioquímica y
Genética
Facultad de las Ciencias de
la Salud
Cra 15 Calle 12 Norte.
Armenia, Quindío
Universidad del Quindío,
Armenia, Colombia
Homero
Sáenz#
Instituto de Errores Innatos
del Metabolismo
Facultad de Ciencias
Pontificia Universidad
Javeriana
Cra. 7 N° 40-62. Bogotá.
D.C., Colombia
Leonardo Lareo†
Grupo de Investigación en
Bioquímica Computacional
Departamento de Bioquímica.
Facultad de Ciencias
Pontificia Universidad
Javeriana
Cra. 7 N° 40-62. Bogotá.
D.C., Colombia
Olga Yaneth Echeverri
Peña
Instituto de Errores Innatos
del Metabolismo
Facultad de Ciencias
Pontificia Universidad
Javeriana
Cra. 7 N° 40-62. Bogotá.
D.C., Colombia
Luis A. Barrera Avellaneda*
Instituto de Errores Innatos
del Metabolismo
Facultad de Ciencias
Pontificia Universidad
Javeriana
Cra. 7 N° 40-62. Bogotá.
D.C., Colombia
E-mail: abarrera@javeriana.edu.co
*Corresponding author
Financial support:
Instituto Colombiano para la Ciencia y la Tecnología Francisco José de Caldas (COLCIENCIAS), Grant No. 1203-12-10410-192-2000 and Pontificia
Universidad Javeriana, Bogotá, D.C., Colombia, Grant No. 120104-O-0101103.
Keywords: E. coli, glycation, human sulfatase, transient expression.
Present address: #Unidad de Biología Celular y Microscopía, Decanato de
Medicina, Universidad Centrocidental Lisandro Alvarado, Ave. Libertador Ave.
Andres Bello, Barquisimeto, Venezuela.
Abbreviations: |
ER: endoplasmic reticulum
hIDS:
human iduronate 2-sulfate sulfatase (native protein)
hrIDS:
human recombinant iduronate 2-sulfate sulfatase
MCB:
master cell bank
MPS
II: hunter syndrome or mucopolysacharidosis type II
OD:
optical density
P(x):
biomass productivity (dry weight)
td: duplication time
WCB:
working cell bank
x:
dry biomass weight
μ(x): Specific growth rate |
The recombinant human iduronate 2-sulfate
sulfatase (hrIDS) was transiently and functionally active expressed in E.
coli K12. The enzyme activity (crude extract) at 100 ml and 400 ml
oscillated between 0.25 and 10.58 nmol h-1 mg-1. The wide
Western-blot peptide profile suggest that hrIDS is proteolitically processed
“randomly” which agrees with the ultrafiltration assay in which the hrIDS
activity was found in all fractions (<30kDa, 30-100kDa and >100kDa). No
glycation sites were found by computer analysis of the hIDS sequence;
discardingthe possibility of marks for glycation and proteolytic processing.
|
