Abstract:
:An approach is presented for the stable covalent immobilization of proteins with a high retention of biological activity. First, chemical modification studies were used to establish enzyme structural and functional properties relevant to the covalent immobilization of an enzyme to agarose based supports. Heparinase was used as a model enzyme in this set of studies. Amine modifications result in 75-100% activity loss, but the effect is moderated by a reduction in the degree of derivatization. N-hydroxysuccinimide, 1,1,1-trifluoroethanesulfonic acid, and epoxide activated agarose were utilized to determine the effect of amine reactive supports on immobilized enzyme activity retention. Cysteine modifications resulted in 25-50% loss in activity, but free cysteines were inaccessible to either immobilized bromoacetyl or p-chloromercuribenzoyl groups. Amine reactive coupling chemistries were therefore utilized for the covalent immobilization of heparinase. Second, to ensure maximal stability of the immobile protein-support linkage, the identification and subsequent elimination of the principal sources of protein detachment were systematically investigated. By using high-performance liquid chromatography (HPLC), electrophoresis, and radiolabeling techniques, the relative contributions of four potential detachment mechanisms-support degradation, proteolytic degradation, desorption of noncovalently bound protein, and bond solvolysis-were quantified. The mechanisms of lysozyme, bovine serum albumin, and heparinase leakage from N-hydroxysuccinimide or 1,1,1-trifluoroethanesulfonic acid activated agarose were elucidated. By use of stringent postimmobilization support wash procedures, noncovalently bound protein loss. An effective postimmobilization washing procedure is presented for the removal of adsorbed protein and the complete elimination of immobilized protein loss.
journal_name
Biotechnol Bioengjournal_title
Biotechnology and bioengineeringauthors
Leckband D,Langer Rdoi
10.1002/bit.260370305subject
Has Abstractpub_date
1991-02-05 00:00:00pages
227-37issue
3eissn
0006-3592issn
1097-0290journal_volume
37pub_type
杂志文章abstract::For the commercial exploitation of lipase biocatalysis to be successful, it is essential that effective supports are selected for lipase immobilization. In this study hydrophobic controlled-pore glasses have been used as model systems for the immobilization of Rhizomucor miehei lipase. The effect of pore diameter and ...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260431006
更新日期:1994-04-25 00:00:00
abstract::A kinetic model describing the cephalosporin biosynthesis in Streptomyces clavuligerus was developed. Using previously reported kinetic data of biosynthetic enzymes, we examined the kinetics of cephalosporin production. The predicted time profile of the specific production rate during a batch culture parallels that of...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260380815
更新日期:1991-10-20 00:00:00
abstract::The feasibility of continuous production of proteins in chemostat cultures of Bacillus subtilis was investigated. An expression system consisting of the bacterium B. subtilis BR151 carrying plasmid p602/19 was used. The plasmid contains the cat (chioramphenicol acetyltrans-ferase) gene downstream of a strong vegetativ...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260470503
更新日期:1995-09-05 00:00:00
abstract::A high-rate anaerobic digestion of food waste leachate were tested using intermittent continuously stirred tank reactors (iCSTRs) to evaluate how severe ammonia inhibition could be mitigated with internal dilution strategy, and to identify how bacterial and archaeal community respond in genus and species level. Experi...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.25968
更新日期:2016-09-01 00:00:00
abstract::Recombinant retroviruses are effective vectors able to integrate transgenes into the target cell's genome to achieve longer-term expression. This study investigates the effect of cell lysis products, a common cell culture by-product, on the transduction of suspension cells by gammaretroviral vectors. Cell lysates deri...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.22634
更新日期:2010-04-15 00:00:00
abstract::Phosphite dehydrogenase represents a new enzymatic system for regenerating reduced nicotinamide cofactors for industrial biocatalysis. We previously engineered a variant of phosphite dehydrogenase with relaxed cofactor specificity and significantly increased activity and stability. Here we performed one round of rando...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.21546
更新日期:2008-02-01 00:00:00
abstract::We describe a method by which the degree of bubble saturation can be determined by measuring the velocity of single bubbles at different heights from the bubble source in pure water containing increasing concentrations of surfactants. The highest rising velocities were measured in pure water. Addition of surfactants c...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260430603
更新日期:1994-03-15 00:00:00
abstract::Monolith reactors combine good mass transfer characteristics with low-pressure drop, the principle factors affecting the cost effectiveness of industrial processes. Recently, these specific features of the monolith reactors have drawn the attention toward the application of the monolith reactor in multiphase reaction ...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.20674
更新日期:2006-02-05 00:00:00
abstract::A two-stage bioreactor scheme was developed for the large-scale production of recombinant proteins using a genetically engineered baculovirus/insect cell system. The first bioreactor was employed for cell growth and the second for cell infection. Silkworm Bm5 cells were infected with a recombinant baculovirus, BmNPV/P...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260420313
更新日期:1993-07-01 00:00:00
abstract::Bacterial biofilms are a major obstacle challenging the development of more effective therapies to treat implant infections. Oxygen availability to bacterial cells has been implicated in biofilm formation and planktonic cell detachment; however, there are insufficient tools available to measure oxygen concentrations w...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.24530
更新日期:2012-10-01 00:00:00
abstract::The knowledge of critical process-relevant genes can be used for an improved control of bioprocesses. So far bioprocess-relevant marker genes can be analyzed by established expression analysis methods only off-line. In this study, an alternative approach for a potential at-line monitoring of gene expression during bio...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.20578
更新日期:2005-11-05 00:00:00
abstract::The rate of celluose degradation, limited due to the inhibition by cellobiose, can be increased by the hydrolysis of cellobiose to glucose using immobilized beta-glucosidase. Production of beta-glucosidase in four yeasts was studied and a maximum activity of 1.22 IU/mg cells was obtained in cells of Pichia etchellsii ...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260260408
更新日期:1984-04-01 00:00:00
abstract::The diffusion of viruses toward cells is a limiting step of the infection process. To be modeled correctly, this step must be evaluated in combination with the adsorption of the virus to the cell surface, which is a rapid but reversible step. In this paper, the recombinant adenovirus (rAd) diffusion and its adsorption...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.21340
更新日期:2007-09-01 00:00:00
abstract::Affinity chromatography using immobilized S-protein was used for the screening of affinity peptide ligands from two soluble peptide libraries. Peptide library I consisted of octamers with glycine (G) at both termini of each peptide, i.e. GXXXXXXG. The six center positions were constructed using random sequences of six...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:
更新日期:1999-06-20 00:00:00
abstract::Ultrafiltration through Carbosep M(4) mineral membrane of protein solutions of decreasing complexity (whey before and after centrifugation or clarification, beta-lactoglobulin) was studied. Mathematical models were used to explain variations in flux with time. Taking into account variations in protein retention and hy...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260340205
更新日期:1989-06-20 00:00:00
abstract::The long hydrocarbon fatty acyl chain is energy rich, making it an ideal precursor for liquid transportation fuels and high-value oleo chemicals. As Saccharomyces cerevisiae has many advantages for industrial production compared to Escherichia coli. Here, we attempted to engineer Saccharomyces cerevisiae for overprodu...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.25239
更新日期:2014-09-01 00:00:00
abstract::An efficient method was developed for media separation and cell collection for eukaryotic cells growing in suspension. The method is based on tangential flow microfiltration using an open channel arrangement in a hollow fiber configuration. Best results (highest processing flux rate) for polysulfone hollow fibers were...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260480412
更新日期:1995-11-20 00:00:00
abstract::Chinese hamster ovary (CHO) cells are the predominant host cell line for the production of biopharmaceuticals, a growing industry currently worth more than $188 billion USD in global sales. CHO cells undergo programmed cell death (apoptosis) following different stresses encountered in cell culture, such as substrate l...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章,评审
doi:10.1002/bit.27269
更新日期:2020-04-01 00:00:00
abstract::A process is described in which the baboon endogenous virus (BaEV) is produced under optimum conditions in cell culture, and concentrated by hollow-fiber ultrafiltration technology under conditions of large-scale production. This system has advantages over conventional systems in that the flow rate is increased 2.5-fo...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260220103
更新日期:1980-01-01 00:00:00
abstract::Mammalian cell cultures typically exhibit an energy inefficient phenotype characterized by the consumption of large quantities of glucose and the concomitant production of large quantities of lactate. Under certain conditions, mammalian cells can switch to a more energy efficient state during which lactate is consumed...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.24728
更新日期:2013-02-01 00:00:00
abstract::A mechanistic model of enzymatic hydrolysis taking into account the morphology of the cellulosic particles and its evolution with time was developed. The individual behavior of the main enzymes involved in the reaction (cellobiohydrolases, endoglucanases, and β-glucosidases), as well as synergy effects, were also incl...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.25873
更新日期:2016-05-01 00:00:00
abstract::Highly efficient antibody immobilization is extremely crucial for the development of high-performance polymeric microdevices for enzyme-linked immunosorbent assay (ELISA). In this article, a site-selective tyrosinase (TR)-catalyzed protein A strategy for antibody immobilization was developed to enhance the sensitivity...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.22136
更新日期:2009-02-15 00:00:00
abstract::Insulin is an important biotherapeutic protein, and it is also a model protein used to study amyloid diseases, such as Alzheimer's and Parkinson's. The preparation of the protein can lead to small amounts of aggregate in the solution, which in turn may lead to irreproducible in vitro results. Using several pre-treatme...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.22902
更新日期:2011-01-01 00:00:00
abstract::A two-stage deterministic model of the growth of Saccharomyces cerevisiae is presented. The cell cycle of this organism was used to suggest the basic model structure. The model represents the preparatory processes of substrate uptake and conversion separately from replication and division. The regulation of the fracti...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260190209
更新日期:1977-02-01 00:00:00
abstract::Among various routes for the biological hydrogen production, the NAD(P)H-dependent pentose phosphate (PP) pathway is the most efficient for the dark fermentation. Few studies, however, have focused on the glucose-6-phosphate 1-dehydrogenase, encoded by zwf, as a key enzyme activating the PP pathway. Although the gluco...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.23259
更新日期:2011-12-01 00:00:00
abstract::To alleviate plasmid instability and to prolong the production phase of subtilisin, integrable plasmid and spore mutants are used. Compared with batch-type shake flask cultures, spore mutants' ability to produce subtilisin can be well pronounced in fed-batch and continuous cultures. Hence, the two culture methods make...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260470610
更新日期:1995-09-20 00:00:00
abstract::Physical damage of animal cells in suspension culture, due to stirring and sparging, is coupled with complex metabolic responses. Nylon microcapsules, therefore, were used as a physical model to study the mechanisms of damage in a stirred bioreactor and in a bubble column. Microcapsule breaskage folowed first-order ki...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260401018
更新日期:1992-12-05 00:00:00
abstract::Exchange of enzyme-bound H(2)O with T(2)O in aqueous solution followed by freeze drying provided tritiated water bound to chymotrypsin, subtilisin Carlsberg, and horseradish peroxidase. The desorption of T(2)O from these enzymes suspended in various organic solvents showed that all three enzymes lost enzyme-bound wate...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260390405
更新日期:1992-02-20 00:00:00
abstract::A biosensor detecting estrogens, progestogens, and androgens in complex samples and in a single step is described. Three Arxula adeninivorans yeast strains were created, each strain producing a different recombinant human hormone receptor and a different fluorescent reporter protein. These strains were then mixed to c...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.26249
更新日期:2017-07-01 00:00:00
abstract::A newly designed batch calorimeter was used to investigate the degradability of some celluloses having varying degrees of crystallinity. The PTC of an aerobic culture of Cellulomonas uda ATCC 21399 obtained revealed a diauxic growth which is attributed to the presence of hemicellulose contaminating Avicel and MN300 ce...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260270711
更新日期:1985-07-01 00:00:00