microbiology, Vol 143, 117-126, Copyright © 1997 by Society for General Microbiology
ScCypB is a novel second cytosolic cyclophilin from Streptomyces chrysomallus which is phylogenetically distant from ScCypA
A Pahl, A Gewies and U Keller
Institut fur Biochemie und Molekulare Biologie, Technische Universitat Berlin, Berlin-Charlottenburg, Germany.
A novel second streptomycete cyclophilin gene-designated sccypB-was
isolated from a cosmid gene library of Streptomyces chrysomallus by using
as gene probe a fragment of the previously isolated cyclophilin gene sccypA
of the same organism. From its sequence the gene sccypB should encode a
protein of M(r) 18868. Expression of sccypB in Escherichia coli as a
hexaHis-tagged fusion protein (H6ScCypB) and enzymic characterization of
the purified protein showed that, like ScCypA, ScCypB is a peptidyl-prolyl
cis-trans isomerase (PPIase). The specific activity and substrate
specificity of the enzyme were comparable to that of ScCypA, but it was
threefold less sensitive to inhibition by cyclosporin A (CsA). In contrast
to ScCypA, which is abundant and exists in free and liganded form, ScCypB
was 50- to 100- fold less abundant in cytosol-derived protein fractions of
S. chrysomallus or Streptomyces lividans, as revealed by Western blot
analyses, suggesting a specialized function for this enzyme in the
streptomycete cell. Both sccypB and sccypA were found to be present as
single copies in the genome of S. chrysomallus and hybridized to a single
band in chromosomal DNAs of other streptomycetes. High-level expression of
sccypB as well as of sccypA cloned into the expression vector pIJ702 did
not produce detectable changes in growth and morphology of S. chrysomallus
and S. lividans. Calculations of similarities to known cyclophilin
sequences and construction of phylogenetic trees indicated that ScCypB and
ScCypA are phylogenetically distant from each other. While ScCypA is
clearly related to the eukaryotic cyclophilins, the analyses show the
sequence of ScCypB to be the most divergent of all cyclophilin sequences,
indicating that it possibly constitutes a cluster by itself.
