Constraints on the transport and glycosylation of recombinant IFN-? in Chinese hamster ovary and insect cells

Hooker, Andrew D.; Green, Nicola; Baines, Anthony J.; Bull, Alan T.; Jenkins, Nigel; Strange, Philip G.; James, David C.

doi:10.1002/(sici)1097-0290(19990605)63:5<559::aid-bit6>3.0.co;2-l

Cited by 109 publications

(68 citation statements)

References 85 publications

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“…It is important to note that this conclusion implies that Sf9 cells can produce and transport CMP-sialic acid, which is required for glycoprotein sialylation in addition to the sialyltransferase activity. This implication is not supported by the results of Hooker et al (1999), who were unable to detect CMP-sialic acid in uninfected or infected Sf9 cells, as discussed above. Based on this finding, the latter group concluded that genetic modification of N-glycan processing in Sf9 cells will be constrained to terminal galactosylation.…”

Section: Engineering Glycoprotein Processing Pathways In the Baculovimentioning

confidence: 73%

“…More recently, Jarvis et al (1998b) constructed several gp64 mutants, each of which had only one of the five potential N-glycosylation sites, and found no evidence of sialic acid in any of the individual N-linked glycans by SNA lectin blotting. Finally, as part of their study, Hooker et al (1999) showed that neither sialyltransferase activity nor CMP-Neu5Ac donor substrate was detectable in baculovirusinfected Sf9 cells, as mentioned above.…”

Section: Glycoprotein Sialylation During Viral Infectionmentioning

confidence: 88%

“…Some investigators have used in vitro assays with radiolabeled or fluorescent CMP-Neu5Ac as the donor substrate to examine insect cell lysates or subcellular fractions for sialyltransferase activities (Butters et al, 1981, Hooker et al, 1999and Lopez et al, 1999.…”

Section: Occurrence Of Endogenous Sialyltransferase Activitymentioning

confidence: 99%

“…However, Hooker et al (1999) recently reported that they were unable to detect any CMP-Neu5Ac by anion exchange chromatography of the soluble nucleotides extracted from uninfected Sf9, Sf21, or Ea4 cells or from baculovirus-infected Sf9 cells.…”

Section: Availability Of Endogenous Cmp-neu5acmentioning

confidence: 99%

“…The N-glycans on interferon γ produced by two insect cell lines (Ea4 and Sf9) were studied by matrix-assisted laser desorption/time of flight (MALDI-TOF) mass spectrometry (Ogonah et al, 1996). Whereas the recombinant protein produced by Sf9 cells had only pauciman-nose-type glycans (see also Hooker et al, 1999), the same protein produced by Ea4 cells contained N-glycans with terminal GlcNAc and Gal residues. However, even the Ea4 cells failed to produce N-glycans containing sialic acid.…”

Section: Use Of Physico-chemical Criteriamentioning

confidence: 99%

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Glycoproteins from Insect Cells: Sialylated or Not?

Marchal

Jarvis²,

Cacan³

et al. 2001

Biological Chemistry

159

122

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Our growing comprehension of the biological roles of glycan moieties has created a clear need for expression systems that can produce mammalian-type glycoproteins. In turn, this has intensified interest in understanding the protein glycosylation pathways of the heterologous hosts that are commonly used for recombinant glycoprotein expression. Among these, insect cells are the most widely used and, particularly in their role as hosts for baculovirus expression vectors, provide a powerful tool for biotechnology. Various studies of the glycosylation patterns of endogenous and recombinant glycoproteins produced by insect cells have revealed a large variety of O-and Nlinked glycan structures and have established that the major processed O-and N-glycan species found on these glycoproteins are (Galβ1,3)GalNAc-O-Ser/Thr and Man3(Fuc)GlcNAc2-N-Asn, respectively. However, the ability or inability of insect cells to synthesize and compartmentalize sialic acids and to produce sialylated glycans remains controversial. This is an important issue because terminal sialic acid residues play diverse biological roles in many glyco-conjugates. While most work indicates that insect cell-derived glycoproteins are not sialylated, some wellcontrolled studies suggest that sialylation can occur. In evaluating this work, it is important to recognize that oligosaccharide structural determination is tedious work, due to the infinite diversity of this class of compounds. Furthermore, there is no universal method of glycan analysis; rather, various strategies and techniques can be used, which provide gly-cobiologists with relatively more or less precise and reliable results. Therefore, it is important to consider the methodology used to assess glycan structures when evaluating these studies. The purpose of this review is to survey the studies that have contributed to our current view of glycoprotein sialylation in insect cell systems, according to the methods used. Possible reasons for the disagreement on this topic in the literature, which include the diverse origins of biological material and experimental artifacts, will be discussed. In the final analysis, it appears that if insect cells have the genetic potential to perform sialylation of glycoproteins, this is a highly specialized function that probably occurs rarely. Thus, the production of sialylated recombinant glycoproteins in the baculovirus-insect cell system will require metabolic engineering efforts to extend the native protein glycosylation pathways of insect cells.

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Section: Engineering Glycoprotein Processing Pathways In the Baculovimentioning

confidence: 73%

Section: Glycoprotein Sialylation During Viral Infectionmentioning

confidence: 88%

Section: Occurrence Of Endogenous Sialyltransferase Activitymentioning

confidence: 99%

Section: Availability Of Endogenous Cmp-neu5acmentioning

confidence: 99%

Section: Use Of Physico-chemical Criteriamentioning

confidence: 99%

See 3 more Smart Citations

Glycoproteins from Insect Cells: Sialylated or Not?

Marchal

Jarvis²,

Cacan³

et al. 2001

Biological Chemistry

159

122

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Choice of Cellular Protein Expression System

Gray¹,

Subramanian

2000

CP Protein Science

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Recombinant protein expression has become a standard laboratory tool, and a wide variety of systems and techniques are now in use. Because there are so many systems to choose from, the investigator has to be careful to use the combination that will give the best results for the protein being studied. This overview unit discusses expression and production choices, including post-translational modifications (e.g., glycosylation, acylation, sulfation, and removal of N-terminal methionine), in vivo and in vitro folding, and influence of downstream elements on expression.

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Protein Glycosylation

Brooks¹

2010

Encyclopedia of Industrial Biotechnology

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More than half of the proteins synthesized by humans are glycosylated. That is, the proteins have one or more N‐ or O‐ linked glycan chains attached to them. Proteins are naturally synthesized in a range of glycoforms, and their glycosylation profile influences their activity, stability, immunogenicity, serum half life, and other biological properties. While the general mechanisms of human protein glycosylation are well established, what influences the fine control of glycosylation patterns is not well understood. Furthermore, the cells of organisms other than humans glycosylate their proteins differently. This is of interest to the biotechnology industry, which commonly uses nonhuman cells for protein expression. Proteins expressed in cells of nonhuman species are glycosylated differently to how they would be by human cells and this is of particular relevance to expression of glycoproteins destined for potential administration to humans. Inappropriate glycosylation profiles result in altered and undesirable pharmokinetic properties. In this chapter, the mechanisms of human protein glycosylation are explained. Glycosylation in cells of nonhuman species, including prokaryotes, fungi and yeasts, insects, plants, and mammals other than humans are introduced, with an emphasis on glycosylation differences of importance to the biotechnology industry. Important advances in engineering glycosylation in nonhuman cell expression systems are highlighted.

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Constraints on the transport and glycosylation of recombinant IFN-? in Chinese hamster ovary and insect cells

Cited by 109 publications

References 85 publications

Glycoproteins from Insect Cells: Sialylated or Not?

Glycoproteins from Insect Cells: Sialylated or Not?

Choice of Cellular Protein Expression System

Protein Glycosylation

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