Expression of C1 Esterase Inhibitor by the Baculovirus Expression Vector System: Preparation, Purification, and Characterization

Wolff, Michael W.; Zhang, Fuming; Roberg, Jeff J.; Caldwell, E.E.O.; Kaul, Patrick R.; Serrahn, Jill N.; Murhammer, David W.; Linhardt, Robert J.; Weiler, John M.

doi:10.1006/prep.2001.1461

Cited by 19 publications

(16 citation statements)

References 31 publications

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“…The expression yields ranged from 3-10 mg/l and, therefore, were comparable to that reported previously for the production of full-length C1 inhibitor using the same expression system (16). In keeping with the report by Wolff et al (16) and in contrast with the highly heterogeneous material produced in P. pastoris (12,19), the C1 inhibitor serpin domain expressed in the current study was homogeneous in terms of size, as shown by SDS-PAGE and MALDI mass spectrometry analyses, indicating relative homogeneity of the N-linked oligosaccharides. In this regard, the mass spectrometry analyses performed on the recombinant proteins are consistent with the occurrence of short, oligomannose carbohydrates comprising two N-acetylglucosamine residues and three to four mannose residues (calculated mass = 893-1055), in keeping with previous analyses (16).…”

Section: Discussionsupporting

confidence: 83%

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Functional Characterization of the Recombinant Human C1 Inhibitor Serpin Domain: Insights into Heparin Binding

Rossi

Bally

Ancelet

et al. 2010

The Journal of Immunology

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Variants of the human C1 inhibitor serpin domain containing three N-linked carbohydrates at positions 216, 231, and 330 (C1inhΔ97), a single carbohydrate at position 330 (C1inhΔ97DM), or no carbohydrate were produced in a baculovirus/insect cells system. An N-terminally His-tagged C1inhΔ97 variant was also produced. Removal of the oligosaccharide at position 330 dramatically decreased expression, precluding further analysis. All other variants were characterized chemically and shown to inhibit C1s activity and C1 activation in the same way as native C1 inhibitor. Likewise, they formed covalent complexes with C1s as shown by SDS-PAGE analysis. C1 inhibitor and its variants inhibited the ability of C1r-like protease to activate C1s, but did not form covalent complexes with this protease. The interaction of C1 inhibitor and its variants with heparin was investigated by surface plasmon resonance, yielding KD values of 16.7 × 10−8 M (C1 inhibitor), 2.3 × 10−8 M (C1inhΔ97), and 3.6 × 10−8 M (C1inhΔ97DM). C1s also bound to heparin, with lower affinity (KD = 108 × 10−8 M). Using the same technique, 50% inhibition of the binding of C1 inhibitor and C1s to heparin was achieved using heparin oligomers containing eight and six saccharide units, respectively. These values roughly correlate with the size of 10 saccharide units yielding half-maximal potentiation of the inhibition of C1s activity by C1 inhibitor, consistent with a “sandwich” mechanism. Using a thermal shift assay, heparin was shown to interact with the C1s serine protease domain and the C1 inhibitor serpin domain, increasing and decreasing their thermal stability, respectively.

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Section: Discussionsupporting

confidence: 83%

“…Functional full-length C1 inhibitor was produced by transient expression in COS-1 cells (15) and using a baculovirus/insect cell system (16). Active C1 inhibitor was also expressed at high yield in Pichia pastoris and purified (17).…”

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confidence: 99%

Functional Characterization of the Recombinant Human C1 Inhibitor Serpin Domain: Insights into Heparin Binding

Rossi

Bally

Ancelet

et al. 2010

The Journal of Immunology

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“…Thus, these results demonstrate that the processing of SEAP N-glycans in both Sf-9 and Tn5Bl-4 cells was inhibited at both low and high DO concentrations. Glycan analysis using CE shows low variability, and is capable of detecting a <10% change in glycan level (Lee et al, 1991;Wol et al, 1999;Choe et al, 2000;Wol et al, 2001). The change observed with varying DO concentration in the most highly processed glycans is approximately 25%, suggesting that the changes are signi®cant.…”

Section: Effect Of Do Concentration On N-glycan Processing In Insect mentioning

confidence: 99%

The effect of dissolved oxygen (DO) concentration on the glycosylation of recombinant protein produced by the insect cell–baculovirus expression system

Zhang

Saarinen

Itle

et al. 2001

Biotech & Bioengineering

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The effect of dissolved oxygen concentration on human secreted alkaline phosphatase (SEAP) glycosylation by the insect cell-baculovirus expression system was investigated in a well-controlled bioreactor. Oligomannose-type N-linked glycans (i.e., Man2 to Man6 and Man3F) were present in SEAP produced by Spodoptera frusiperda Sf-9 (Sf-9) and Trichoplusia ni BTI-Tn-5B1-4 (Tn-5B1-4) insect cell lines. The relative amounts of the most highly processed glycans (i.e., Man3F and Man2 in the SEAP from Sf-9 and Tn-5B1-4 cells, respectively) were significantly higher at 50% of air saturation than at either 10% or 190% of air saturation. That is, glycan processing was inhibited at both low and high dissolved oxygen concentrations.

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“…Recombinant C1 inhibitor (Pharming Group NV, Leiden, Netherlands) has been developed and trials should be available in the near future [94].…”

Section: Potential New Therapiesmentioning

confidence: 99%

C1 inhibitor deficiency: consensus document

Gompels

Lock

Abinun

et al. 2005

Clinical and Experimental Immunology

379

460

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SummaryWe present a consensus document on the diagnosis and management of C1 inhibitor deficiency, a syndrome characterized clinically by recurrent episodes of angio-oedema. In hereditary angio-oedema, a rare autosomal dominant condition, C1 inhibitor function is reduced due to impaired transcription or production of non-functional protein. The diagnosis is confirmed by the presence of a low serum C4 and absent or greatly reduced C1 inhibitor level or function. The condition can cause fatal laryngeal oedema and features indistinguishable from gastrointestinal tract obstruction. Attacks can be precipitated by trauma, infection and other stimulants. Treatment is graded according to response and the clinical site of swelling. Acute treatment for severe attack is by infusion of C1 inhibitor concentrate and for minor attack attenuated androgens and/or tranexamic acid. Prophylactic treatment is by attenuated androgens and/or tranexamic acid. There are a number of new products in trial, including genetically engineered C1 esterase inhibitor, kallikrein inhibitor and bradykinin B2 receptor antagonist. Individual sections provide special advice with respect to diagnosis, management (prophylaxis and emergency care), special situations (childhood, pregnancy, contraception, travel and dental care) and service specification.

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Expression of C1 Esterase Inhibitor by the Baculovirus Expression Vector System: Preparation, Purification, and Characterization

Cited by 19 publications

References 31 publications

Functional Characterization of the Recombinant Human C1 Inhibitor Serpin Domain: Insights into Heparin Binding

Functional Characterization of the Recombinant Human C1 Inhibitor Serpin Domain: Insights into Heparin Binding

The effect of dissolved oxygen (DO) concentration on the glycosylation of recombinant protein produced by the insect cell–baculovirus expression system

C1 inhibitor deficiency: consensus document

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