Properties of PBCV WAL (wall associated) lysin
Properties of PBCV WAL (wall associated) lysin
Hypothesis was lysozyme-like enzyme which bound to cell walls
Original purification protocol (dissociate from "wall fraction" (cell residue) with high salt)
(compare isolation procedure to that of ribosomal initiation factors)
WAL lysin acts on cells of all endosymbiotic Chlorella (and a few others), but not on all Chlorella
This contrasts with intact virions which are host specific
pH optimum around neutral
Can be purified and stored @ pH 4.5
Requires moderate salt concentration
Stable in the fridge (months), loses activity above 50C
Stimulated by divalent cations, calcium and strontium work well
Resistant to PMSF, DTT, IAm, hydrogen peroxide, borate, bisulfite
Inhibited by polycations, sarkosyl, EGTA, diethyl pyrocarbonate, proteases, antibody to PBCV
Fails to hydrolyze a collection of umbelliferyl sugars
Alginate lyase is the only commercial enzyme with (weak) lysin activity
Lysin doesn't hydrolyze alginate (Chlorella walls are the only lysin substrate)
Salt activates lysin in PBCV virions - viscosity indicates some DNA release
Conditions (high salt) which activate lysin in virions resemble those which extract lysin from "wall".
Protein composition of lysin preps ("WAL lysin" enriched in virions, not in cell residue)
Native gels of lysin preps
Gel diffusion assay of lysin - Overlaying native gels (above) on Chlorella agarose shows no digestion
Protamine sulfate effectively removes DNA from lysin preps
Gel filtation of lysin prep (comes off in void volume of G200)
Gel filtration of most recent prep
Lysin components do not bind detectably to cell walls