Fractionation of a whey growth factor for Streptococcus agalactiae into two active components containing proteins

Previously three factors (F-1, F-2, F-3) that stimulated in vitro growth of Streptococcus agalactiae were separated from wheys of milk and colostrum by chromatographic procedures. Now F-1 was separated further into two active components, F-1a and F-1b, when rechromatographed on an anion exchange resin with distilled water as eluent. The stimulatory activity was associated with two protein peaks whereas a third protein peak that was eluted in those fractions containing lactose was not stimulatory. Most of the stimulatory activity (76 to 80%) and protein (90 to 93%) were retained by a filter with retention of greater than 1,000 MW. Two stimulatory factors were confirmed by separation of F-1 fractions by step-gradient elution on a column of octadecylsilica; one protein peak was found for each of the three solvents. Samples with protein eluted with .1 N formic acid and formic acid: methanol (1:1) had stimulatory activity, whereas the sample with protein eluted with 100% methanol was inactive. Results were similar when F-1 fractions were applied to Sep-Pak (muBondapack C18) cartridges and eluted with water, 50% methanol, and 100% methanol. Proteins in the Sep-Pak eluates with stimulatory activity (water and 50% methanol) were heterogeneous and different when separated by high-voltage paper electrophoresis. Proteose-peptone preparations with added lactose were chromatographed on columns of anion exchange resin, with distilled water as eluent, and produced two peaks of protein with elution volumes comparable to peaks 1 and 3. Lactose appeared to be a major factor in eluting the second protein peak.(ABSTRACT TRUNCATED AT 250 WORDS)