Recombinant glycoproteins expression in insect cell....

[Friedrich Altmann]

The question whether insect cells do sialylate and if so, when and 
what, is indeed a rather hot topic. The situation is not quite simple.
There are a few reports claiming sialylation of O- or N- or something-
glycans. 
Roth et al. (Science 256, p. 673 found a polysialylated protein 
in Drosophila larvae. 
Vandenbroeck et al (Lymphokine Cytokine Res. 13, p. 253..) found 
complex N-glycans. Sridhar et al. (FEBS Lett. 315, p. 282.) and Hafer 
and Ferenz (Comp. Biochem. Physiol. 100B, p. 579..) detected sialic 
acid on their recombinant glycoproteins. These three studies were done 
with lectins and/or neuraminidase and/or enzymatic determination of
sialic acid. Such methods have their merits, however, they must be 
rated as unreliable to some extent.
And there are the studies of Dr. Castellino and coworkers mentioned by
R. Schnaar. They have in fact received a lot of attention and might 
well be citation classics in the field. There are however a few 
problems with his results.

1.) For at least four years now, nobody else could confirm these 
results - although a lot of people tried.

2.) To my knowledge, Castellino has not undertaken to directly 
communicate with other researchers in the field on this topic. Does he 
know why all the others did not ´succeed´ ? Does he use another 
baculovirus or a different cell line than all the others ?
I would very much appreciate to hear or read his current opinion.   

3.) My limited knowledge of cell biology does not provide me with an 
idea how the expression of recombinant plasminogen might influence the
expression of non-transcribed or non-translated proteins. Anyway, all 
other baculovirus constructs failed to effect such changes of genetic 
regulation.

Therefore, the nasty suspicion might arise that there´s something
wrong with Castellino´s studies. He is cordially invited to dissipate
such doubts.

Apart from that I want to suggest that whenever a sialylated insect
glycoprotein seems to appear, check it´s identy, don´t rely on 
lectin blots or enzymatic methods alone. Cooperate with groups
experienced in glycan analysis (to make it clear: we are no sialylation 
experts) and do not rely on commercial kits (which are like tarts: 
expensive but you often won´t get what you want).  
_____________________________________________________________________

Altmann Friedrich
Universtitaet fuer Bodenkultur Vienna
A-1180 Austria
email: altmann at edv1.boku.ac.at