Deoxyribonuclease of Syncephalastrum racemosum--enzymatic properties and molecular structure.
Among the isolated fungal species of soil, one filamentous fungus, Syncephalastrum racemosum, produces a relatively large amount of DNase. This enzyme has been purified to apparent homogeneity by column chromatography on DEAE-cellulose, hydroxylapatite, phenyl-Sepharose CL-4B, and Sephadex G-100. The active enzyme requires divalent metal ions and has an optimum pH of 7.0 with Mg2+ and 7.2 with Mn2+. This enzyme is an acidic glycoprotein with a pI 5.0 and is relatively unstable at low concentrations. The M(r) of the enzyme is 56,000 during gel filtration under nondenaturing conditions but is 28,000 during polyacrylamide gel electrophoresis in sodium dodecyl sulfate. These results suggest a structure consisting of two subunits. The subunits of the holoenzyme can be cross-linked with glutaraldehyde. The yield of N-terminal phenylthiohydantoin-alanine from the holoenzyme is 140% and that of one peptide (D-Y-V-S-S-G-Y-D-R), obtained from the tryptic digest is 160%, indicating that the native enzyme is composed of two identical subunits and probably has two active domains. Fungal DNase can be inactivated by Cu(2+)-iodoacetate under conditions that inactivate bovine pancreatic DNase. The specific activity (units/mg of protein) of fungal DNase is 6.5 times that of bovine DNase. The amino acid content of fungal DNase, relative to bovine DNase, is higher in Gly and lower in Ser and Val. The fungal N-terminal 40-residue sequence shows a high degree of homology with a consensus sequence derived from DNase of three mammalian species.
Chen
L Y
LY
Institute of Biochemistry, College of Medicine, National Taiwan University, Taipei, Republic of China.
Ho
H C
HC
Tsai
Y C
YC
Liao
T H
TH
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D002845:Chromatography; D003851:Deoxyribonucleases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D005656:Fungal Proteins / Q000737:chemistry; D005658:Fungi / Q000201:enzymology*; D005976:Glutaral; D006023:Glycoproteins / Q000737:chemistry; D007461:Iodoacetates / Q000494:pharmacology; D019807:Iodoacetic Acid; D008969:Molecular Sequence Data; D015394:Molecular Structure; D012988:Soil Microbiology; D013057:Spectrum Analysis
D000596:Amino Acids; D005656:Fungal Proteins; D006023:Glycoproteins; D007461:Iodoacetates; D003851:Deoxyribonucleases; D005976:Glutaral; D019807:Iodoacetic Acid