Generally speaking, there is a lack of heavy chains with which 2m can pair, and so the excess 2m in the US11+

cells behave as a secretary protein would. Due to this, they can be compared to control cells by comparing concentrations found in recovered supernatants. The light chain of the MHC Class I molecule has been shown to be very stable through the results of various experiments. The 2ms behaviour is not affected by the inclusion of the inhibitor LLnL, as the inhibitor has no observable effect on the actual quantity of 2m released. If LLnL induces the breakdown of the heavier, Nglycosylated chain into its 40 kDa intermediate, the intermediate will not form an association with the 2m chain even though this association in biosynthesis is a very rapid process as no detectable reduction of 2m secretion has been detected. Therefore, to maintain the lighter 2m chains stability, it seems that the US11+ gene decreases the likelihood of the association of the lighter chain and the heavier chains intermediate. When the breakdown of the heavier chains 40 kDa intermediate was first observed in US11+ cells treated with LLnL, two potential theories were put forward as to how it happened. It was first suggested that some form of proteolytic cleaving removed the cytoplasmic tail of the intermediate. However, antibodies obtained from the cytoplasmic COOH terminus of MHC Class I molecules are still able to recognise the breakdown of the intermediate; this was indicative of the intermediates COOH terminus still remaining, mostly, intact. It was next suggested that the heavy chains themselves were being modified and so pulse-chase experiments were carried out using the enzyme endoglycosidase H, or endo H. This enzymes operates by cleaving the high mannose-type oligosaccharides which are commonly found on ER-resident N-linked glycoproteins. The figure above, taken from the article, shows the results of an experiment to demonstrate the recovery of MHC Class I proteins using the W6/32 antibody in control cells, US11+ cells and cells which were exposed to LLnL. When both the graphs, we can see that the intermediate of the heavier chains in US11+ cells is almost fully resistant to Endoglycosidase H once treated with BFA. However, it is only detected once the cells were exposed to LLnL, although the levels are still somewhat lower than those in the control. Also, the mobility of the intermediate is indistinguishable from the normal, control cells, which led the researchers to conclude that the breakdown product of the intermediate contains very little, if any, N-linked glycans.