Purification of GTP:alpha-D-mannose-1-phosphate guanyltransferase.

The enzyme GTP:alpha-D-mannose-1-phosphate guanylyltransferase from porcine thyroid tissue has been purified 69 900-fold on columns of blue-Sepharose, DEAE-Sepharose, phenyl-Sepharose and agarose-GTP affinity materials. Although it exhibits a tendency to aggregate, the enzyme travelled, upon sucrose velocity sedimentation, as a single oligomer with a molecular mass of 412 kDa. Michaelis constants were determined to be 1.0 microM, 1.0 mM, 3.5 microM and 0.4 microM for GDP-alpha-D-mannose, pyrophosphate, GTP and mannose-1-phosphate, respectively. The enzyme appears to be specific for the mannose moiety but will accept an inosine replacement for guanine and a deoxyribose replacement for ribose in GTP.