Glycoprotein biosynthesis: lipid intermediates Frank H E M M I N G
Department of Biochemistry, University of Nottingham, Queen's Medical Centre, Nottingham NG~ 2U, U.K. Rather than attempt t.o summarise all the good posters and talks, which would probably be unnecessary and would certainly take a long time, I thought it might be better to restrict myself to some general impressions. Some of these have been strengthened and some have been weakened during the week. As a teacher, I try to identify generalisations whereby I can try to transmit ideas to students who I like to think might get interested in our area of glycoconjugate research. One of the generalisations that I am in the habit of making has been weakened somewhat and clearly now needs modifying. This concerns the dogma that, in glycosyl transfer using dolichyl phosphate, oligosaccharides are always linked through a diphosphate and monosaccharides through a monophosphate. Well earfier in the week Professor Wieland told us that the Halobacteria do it slightly differently in that they link oligosaccharides to dolichol v/a a monophosphate. I wonder if this is the only exception to the dogma. One of my main interests in the biosynthetic area concerns those factors that control or can be used to manipulate the glycosyiation of proteins, i suppose that this has coloured my thinking in preparing for this afternoon. I get the impression that we are edging our way at leas*, to getting the methodology sorted out for making real progress in the use of molecular biology in this area. Gene and molecular cloning are clearly very powerful techniques and we saw some applications this morning. Although compared with the promise 2 years ago, I had hoped for rather more news of cloning successes in a few other areas this year. If we are thinking of the control of biosynthesis, we do have to concern ourselves with genetic control, probably of several different types of proteins: transferases, glycosidases and transporters. Perhaps we can hope that in 2 or 4 years time we will have a clearer pic'~are of the common features of those proteins that bind similar compounds. Just as we heard thi:~ morning of some features repeated among the ga~actose-binding proteins, we can expect to find aspecB of protein structure recurring among, for example, ~:hoseenzymes using dolichyl phosphate as a result of cloning the transferases. Of course we presume that many of these enzymes work as parts of multi-enzymes complexes which are concerned with the overall elaboration of oligosaccharides. We have enzymes, transporters and possibly proteins that help to hold the complex together. That clearly
is an area in which we await progress with great interest. But the genetics is not the only aspect of control that is important. Physiological control is also of interest and we heard today of cAMP-dependent phosphorylation having some effect in the parotid gland on dolichyl phosphate synthetase activity. It will be very interesting to know how general this phenomenon is both with respect to other cell types and to other enzymes. We also heard about cAMP switching on gene expression of proline-rich proteins. There was from Professor Nagai just a hint that there may be other receptor-mediated processes involving growth factors that may influence the activity of, or the expression of, enzymes concerned with the synthesis of oligosaccharides of glycolipids. Also very important is the lipid environment and the way in which the enzymes are located in the membrane. I was fascinated by Dr. Hirschberg's talk on the topographical distribution of enzymes in the rough endoplasmic reticulum that use dolichyl phosphate. I wonder if in fact there may be some activities on the cytoplasmic side of the membrane and, linking these thoughts with other lectures, if it may be possible to titrate out some of these activities using inhibitors of the type that Dr. Schwarz told us about this morning. I wonder also if insistence on interpreting the results of this type of experiment always in terms of the bilayer may not sometimes be a mistake. There is little doubt that the bilayer is the best description of most biological membranes especially the plasma membrane but there is increasing evidence for the presence of H-2 phase membrane conformations and it is particulary interesting that Dr. Schutzbach has reminded us again at this meeting that several of the enzymes of the dolichyl phosphate pathway much prefer an environment of phospholipids and other lipids that favour that sort of arrangement rather than the bilayer. It may well be that where we get discontinuities of the membrane - say underneath the ribosome or at the budding off of vesicles, remembenng that membranes are very dynamic structures we will find phenomena that are more accurately described in terms of H-2 phase. Finally, together with several of the other chairmen, I was intrigued to hear of the progress being made with the Pl-anchor and the sort of problems that it presents for someone interested in biosynthesis. Is this going to be another pathway that involves doli¢hyi phosphate mannose?