Isolation of a human serum protein that inhibits the growth of Cryptococcus neoformans.
Human serum at 5 to 10% (v/v) in tissue culture medium RPMI-1640, inhibits the growth of Cryptococcus neoformans by 80 to 93%. Serum fractionated on molecular sieve columns (Sephadex G-200) yielded an active protein fraction. This fraction at 100 micrograms protein/ml inhibited the growth of C. neoformans by 54%. When an active G-200 fraction was applied to a dye affinity column (Affi-Gel Blue) the fraction with inhibitory activity was bound by the column and was eluted with 1.4 M NaCl in 0.1 M phosphate buffer (pH 7.4). The bound fraction at 62.5 micrograms protein/ml inhibited C. neoformans growth by 82%. On native polyacrylamide gel electrophoresis (Nu-PAGE) the bound fraction migrated as a major and a minor band. Under the reducing conditions of sodium dodecyl sulfate (SDS)-PAGE the bound fraction yielded 4 prominent bands with MW ranging from 175 kDa to 45 kDa. Purification of the active Sephadex G-200 peak was achieved using an anion exchange column (DEAE-Sephacel). Protein eluted with 0.1 M NaCl had strong anticryptococcal activity (12.5 micrograms/ml, 79% inhibition), which in SDS-PAGE migrated as a single band with an approximate MW of 85 kDA. This protein appears important in natural host resistance to C. neoformans and polymorphisms or deficiencies may have epidemiologic and diagnostic relevance.
Duvvuru
S
S
California Institute for Medical Research, San Jose, USA.
Brummer
E
E
Morelli
R
R
Stevens
D A
DA
D000935:Antifungal Agents / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D001798:Blood Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology*; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003455:Cryptococcus neoformans / Q000187:drug effects* / Q000254:growth & development; D003470:Culture Media; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans
D000935:Antifungal Agents; D001798:Blood Proteins; D003470:Culture Media