M. Beukes et al., Purification and partial characterization of a murein hydrolase, millericin B, produced by Streptococcus milleri NMSCC 061, APPL ENVIR, 66(1), 2000, pp. 23-28
Streptococcus milleri NMSCC 061 was screened for antimicrobial substances a
nd shown to produce a bacteriolytic cell wall hydrolase, termed millericin
B, The enzyme was purified to homogeneity by a four-step purification proce
dure that consisted of ammonium sulfate precipitation followed by gel filtr
ation, ultrafiltration, and ion-exchange chromatography. The yield followin
g ion-exchange chromatography was 6.4%, with a greater-than-2,000-fold incr
ease in specific activity, The molecular weight of the enzyme was 28,924 as
determined by electrospray mass spectrometry, The amino acid sequences of
both the N terminus of the enzyme (NH2 SENDFSLAMVSN) and an internal fragme
nt which was generated by cyanogen bromide cleavage (NH2 SIQTNAPWGL) were d
etermined by automated Edman degradation. Millericin B displayed a broad sp
ectrum of activity against gram-positive bacteria but was not active agains
t Bacillus subtilis W23 or Escherichia coli ATCC 486 or against the produce
r strain itself, N-Dinitrophenyl derivatization and hydrazine hydrolysis of
free amino and free carboxyl groups liberated from peptidoglycan digested
with millericin B follow ed by thin-layer chromatography shelved millericin
B to be an endopeptidase with multiple activities. It cleaves the stem pep
tide at the N terminus of glutamic acid as well as the N terminus of the la
st residue in the interpeptide cross-link of susceptible strains.