Master of Science (MS)
Semester of Degree Completion
William A. Weiler
The isolation of Rhizobium japonicum cell walls was performed by ultrasonication and the enzymatic digestion with DNase, RNase and pronase separately. Crude cell walls were washed with M NaCl and 5% SDS and were isolated by 20,000 g centrifugation. Muropeptide amino acids were freed by hydrolysis at 121C for 10 minutes in 6N HCl.
Detection of Rhizobium japonicum murein layer amino acids was done by 2-dimensional instant thin layer chromatography on silica gel plates. The solvents found most favorable, were n-butanol: acetic acid: water and n-propanol: 34% ammonium hydroxide. Amino acid spots were developed by spraying ninhydrin on the chromatograms, followed by heating at 100C for 10 minutes. Spots were identified by their Rf values compared to those of known amino acids run in the same solvent system.
The eight amino acids and sugars identified in the murein layer of Rhizobium japonicum cell walls were DAPA, lysine, alanine, glutamic acid, glucosamine, serine, (valine, methionine or isoleucine) and leucine.
Alauddin, Mohammed, "Studies on Muropeptide Amino Acids of Rhizobium japonicum" (1973). Masters Theses. 3705.