A 29 kb shuttle cosmid vector, pTYS507, was constructed from a cryptic Micromonospora griseorubida plasmid and the Escherichia coli cosmid pJB8. Subcloning of mycinamicin II biosynthesis genes in pTYS507 led to the identification of a DNA region that could complement a mutant of M. griseorubida that lacked both hydroxylase and epoxidase activities. Nucleotide sequence and mutational analysis suggested that a single P-450-like protein catalyzes both reactions.
Nanofiltration technology to remove possible pathogenic viruses during biopharmaceutical
manufacturing was introduced in the biopharmaceutical industry in 1989. The very first industrial
implementation took place in the early 1990s, through commercial manufacturing processes of plasma-
derived medical products. Then it was applied to recombinant protein medical products, including
monoclonal antibodies. In the first review published in 2005 in this journal, the technology was
already considered promising and was much welcomed by the industry, but it was still a relatively
emerging technology at that time, and many questions were raised about its robustness as a reliable
virus-removal tool. We conducted a review to update the published information (SCI journals and
suppliers’ documentation) existing on the use of nanofiltration as an industrial process for removing
viruses from various biologicals. After almost a decade from the previous review, nanofiltration has
established itself as a routine production step in most biopharmaceutical manufacturing. It has become
one of the essential manufacturing processes used to assure safety against viral contamination.
The technology is applied to manufacturing processes of various biologicals (human plasma products
and complex recombinant proteins, such as coagulation factors and monoclonal antibodies made
from mammalian cells). Many biologicals that undergo nanofiltration are licensed by regulatory
authorities, which illustrates that nanofiltration is recognized as a robust and safe virus-removal method.
No adverse events related to the use of nanofiltration have been recorded. New trends in
nanofiltration technology continue to appear. As was identified during its introduction to the market
and predicted in the previous review, nanofiltration has achieved major technical breakthroughs for
ensuring the safety of biologicals, particularly human plasma-derived products, against viruses.
Micromonosporastrains produce some important antibiotics, such as gentamicins1*, sisomicins2), sagamicins3), fortimicin A4) and mycinamicins5). We discovered mycinamicins, 16-membered macrolide antibiotics with a strong antimicrobial activity against Gram-positive bacteria, and reported biosynthetic pathway of mycinamicin II6). It has been our interest to clarify the mechanismsof regulation and expression of the genes for mycinamicin biosynthesis in Micromonospora griseorubida in order to improve the producing strains. For this purpose, it was necessary to establish a system to express the cloned genes in M. griseorubida. hours. Protoplasts were filtered through cotton wool, centrifuged and resuspended in 1ml of P medium15). To obtain a good yield of protoplasts, growth phase turned out to be effective as pointed out in Streptomyces16). A dramatic increase in yield was observed using mycelia for 44 hours incubation (early stationary phase). While the number of protoplasts obtained from 36 hours incubated culture was 108/ml, that from 44hours culture provided 1010/ml.For regeneration of M. griseorubida protoplasts, R2YE17) generally used for regeneration of Streptomyces protoplasts, gave the regeneration frequency as low as < 0.1 %. Therefore, we designed MR0.3S by modifying the M agar, which was used for the cultivation of M. griseorubida. MR0.3S medium contained sucrose 103.0 g, soluble starch 20.0 g, soy bean meal 7.5 g, Polypepton 5.0 g, CaCO3 1.0 g, MgSO4 à" 7H2O 5.0 g, trace elements solution15) 10.0ml, FeSO4-7H2O 40.0mg and Bacto-agar 22.0g, per liter of deionized water (adjusted to pH 7.2 with 1 n NaOH
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.