Subunit composition of a high molecular weight oligomer: Limulus polyphemus hemocyanin

The hemocyanin of Limulus polyphemus is a 48-subunit aggregate. This 3.3 × 10⁶-dalton oligomer is composed of structurally and functionally heterogeneous subunits. Using polyacrylamide electrophoresis J. Markl, A. Markl, W. Schartau, and B. Linzen (J. Comp. Physiol. Ser. B130,283–292, 1979) observed 12 bands; while using immunoelectrophoresis, M. Hoylaerts, G. Preaux, R. Witters, and R. Lontie (Arch. Int. Physiol. Biochem.87, 417–418, 1979) and J. Lamy, J. Lamy, J. Weill, J. Bonaventura, C. Bonaventura, and M. Brenowitz. (Arch. Biochem. Biophys.196, 324–339, 1979) observed 8 subunits. To proceed with an analysis of subunit roles in assembly it is first necessary to determine the number of distinct subunits. Refinement of the chromatographic separation procedures has led to the isolation of 8 immunologically distinct subunits as well as additional charge isomers which cannot be distinguished immunologically. Alkaline electrophoresis revealed 15 bands and isoelectric focusing up to 17. On the basis of extensive control experiments, including composit acrylamide-agarose immunoelectrophoresis and checks for conformational isomers, aggregation, proteolysis, and other types of degradation, we conclude that the electrophoretic heterogeneity of immunologically identical subunits is not artifactual. We have extended the nomenclature used by Lamy et al. (1979) to include the electrophoretic heterogeneity by using primes (′) to denote electrophoretically distinguishable subunits which are immunologically identical. A number of patterns have become apparent by correlating the results obtained by the different techniques. For example, immunologically pure subunit II, which shows 3 bands on alkaline electrophoresis, is in fact a mixture of electrophoretically distinct subunits II, II′, II″. Except for subunits II, II′, and II″ immunoelectrophoretically identical subunits are typically homogeneous on sodium dodecyl sulfate-gels. However, slight differences in the apparent molecular weight are observed on high-resolution gels between immunologically unrelated subunits. The immunological identity and electrophoretic differences suggest that the charge isomers which are immunologically identical have similar antigenic surfaces. If a charge substitution is not in a critical location, we would expect the electrophoretically distinct but immunologically identical subunits to have identical assembly roles. Comparison of the results for Limulus hemocyanin with the hemocyanin of related species Eurypelma californicum and Androctanus australis, which have 7 and 8 immunologically distinct subunits, respectively, suggests that the calcium-mediated aggregation from 24 to 48 subunits of Limulus does not require more extensive subunit complexity.     

Epithelial impedance analysis in experimentally induced colon cancer.

Epithelial impedance analysis was used to measure the alterations in resistance of the large bowel in a murine model of large bowel cancer. The technique was able to resolve the epithelial resistance from the total resistance of the bowel wall. A progressive decrease in resistance of the bowel epithelium occurs during carcinogenesis induced with dimethyhydrazine. About a 21% decrease in epithelial resistance from 22.0 +/- 1.3 omega.cm-2 to 17.5 +/- 1.1 omega cm-2 (p less than 0.025) was observed after 20 wk of carcinogen administration. The sensitivity of the technique in detecting altered epithelial resistance in premalignant bowel mucosa was improved by examining the impedance profile in a sodium-free Ringer's solution where the epithelium of control colons had a resistance of 24.4 +/- 1.8 omega.cm-2 compared with 19.0 +/- 1.1 omega.cm-2 (p less than 0.02) in colons from animals treated for only 4 wk with the carcinogen. Epithelial impedance analysis would seem to be a sensitive technique capable of identifying changes in the electrical properties or the large bowel early in disease states.     

Book Review

Book Reviewin this Article:
Diabetic Neuropathy edited by P. J. Dyck, P. K. Thomas, A. K. Asbury, A. I. Winegrad, and D. Porte, Jr.