• 한국미생물·생명공학회지
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• 제17권6호
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• pp.611-618
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• 1989

Cephalosporium acremonium 균주로 미생물 고정화 증식입자를 제조하여 유동층 생물반응기에서 세파로스포린 C 생산에 대한 메치오닌의 영향에 대하여 연구하였다. 메치오닌은 세파로스포린 C 생합성에 매우 중요한 대사조절인자로 알려져 있는데, 본고에서는 초기 발효배지 내 메치오닌 농도의 영향과 이를 토대로 생물반응기에 메치오닌을 주입하였을 때 세파로스포린 C 생산에 미치는 메치오닌의 영향을 관찰하였다. 초기 배지 내에 존재하는 메치오닌에 의해 세파로스포린 C 생산성이 증가하였으며, 메치오닌의 최적양(0.3-0.5w/v％)이 존재하였다. 또한 메치오닌에 의해 탄소원의 소모속도가 증가됨이 관찰되었다. 배지내 메치오닌이 고갈될 때 세파로스포린 C 생산이 극대화되었으나 유동층 생물반응기에서 메치오닌을 부가적으로 첨가하였을 때 그 효과는 기대한 수준에 미치지 못했는데 이는 생체내 메치오닌의 축적과 관계가 있는 것으로 판단되었다. 또 고정화 증식입자를 사용하였을 때 세파로스포린 C 생산에 양호한 결과를 얻었는데 이 경우에도 적정량의 메치오닌 투여가 생산성에 중요한 인자임을 발견하였다. 따라서 유동층 반응기의 운전에 의해 세포로스포린 C 생산성을 향상시키기 위해서는 초기 운전단계에서 최적양의 메치오닌을 주입하는 것이 가장 중요하며 고정화 증식입자의 사용에 의해 공정생산성을 획기적으로 증대시킬 수 있다고 판단되었다.

• KSBB Journal
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• 제10권3호
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• pp.264-270
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• 1995

Trigonopsis variabilis는 버l타락탐 항생제인 cephalosporin C (ceph C)를 ${\alpha}$-keto-adipyl-7 a aminocephalosporanic acid(AKA-7 ACA)로 생변 환하는 강력한 D-amino acid oxidase 효소를 갖고 있다. 본 연구는 이 D-AAO 효소의 유전자를 추출하기 위하여 polymerase chain reaction (PCR)을 사용하였다. PCR 단편을 콜로닝하기 위하여 Taq D DNA polymerase, Klenow, T4 DNA polymerase I, Alkaline phosphatase Calf Intestinal와 T4 kinase 의 효소반응을 이용하여 4가지의 방법을 샤용한 결 과, blunt - end 의 PCR fragment 를 phosphory­l lation하고 blunt -end의 pUC18 plasmid를 dephos phorylation 한 후 ligation 한 것 이 가장 좋은 클로 닝 효율을 보였다. Ceph C에 대한 D-AAO 효소의 활성은 재조합 E. coli의 세포추출액과 permea bilized cells에서 모두 확인할 수 있었다.

• Journal of Microbiology
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• 제37권4호
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• pp.200-205
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• 1999

7-Aminocephalosporanic acid (7-ACA) is the initial compound in preparation of cephalosporin antibiotics widely used in clinical treatment. Bacteria producing glutaryl 7-ACA acylase, which convert cephalosporin C to 7-ACA, has been screened in soil samples. A bacterial strain exhibiting high glutaryl 7-ACA acylase activity, designated KAC-1, was isolated and identified as a strain of Pseudomonas diminuta by characterizing its morphological and physiological properties. The screening procedures include culturing on enrichment media containing glutaric acid, glutamate, and glutaryl 7-aminocephalosporanic acid as selective carbon sources. To enhance enzyme production, optimal cultivation conditions were investigated. This strain grew optimally at pH 7 to 9 and in temperatures of 20 to 40 C, but acylase production was higher when the strain was grown at 25 C. Glutaric acid, glutamate and glucos also acted as inducers for acylase production. In a jar fermenter culture, P. diminuta KAC-1 produce acylase in a growth-associated manner. The substrate specificity of KAC-1 acylase by cell extract showed that this enzyme had specificity toward glutaryl 7-ACA, glutaryl 7-ADCA, but not cephalosporin C.

• Archives of Pharmacal Research
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• 제15권3호
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• pp.234-238
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• 1992

Using cell-free extract of Lysobacter lactamgenus, enzymatic conversion of $\delta$-L-($\alpha$-aminoadiphyl)-L-cysteinyl-D-valine (ACV) the first substrate of $\beta$-lactam biosynthesis, into antibiotic compounds was attempted. In high performance liquid chromatographic (HPLC) analysis, the biosynthetic intermediates for cephalosporin antibiotics including isopenicillin N, deacetoxycephalosporin C, deacetylcephalosporin C and unknown cephem compound were detected in reaction mixtures. It implies that cephabacin compounds from L lactamgenus could be produced by biosynthetic routes through penicillin ring formation and its expansion to cephalosporin ring, likely as cephalosporin C from Cephalosporium or cephamycin C from Streptomyces. Among biosynthetic enzyme in cell-free extract, the ring formation activity (isopenicillin N synthetase activity) was separated in 50-60% of ammonium sulfate fraction, and ring expansion activity (deacetoxycephalosporin C synthetase activity) was found to be in 40-50% fraction. The partially purified isopenicillin N synthetase could convert as much as 90% ACV to isopenicillin N during 6-hour reaction.

• Biotechnology and Bioprocess Engineering:BBE
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• 제6권2호
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• pp.156-156
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• 2001

The objective of this study was to investigate the effects of nutrients and culture conditions on morphology during the seed culture of C. acremonium ATCC 20339 Morphological factors such as hyphal length number of tips number of arthrospores were observed to investigate the relationship between seed morphology and CPC production. During the time course of seed culture, hyphal length was shortened and the number of arthrospores increased rapidly On the other hand the number of tips deceased rapidly and this was closely related to the hyphal length Mixed nitrogen sources of 3% solybean meal and 1% cotton seed flour were determined as the proper organic nitrogen sources, in terms of the morphological factors in the seed culture. This fact was proven in batch culture for the production of Cephalosporin C. It was also found that a proper agitation speed enhanced the morphological differentiation of C. acremonium ATCC 20339, thus improving the production of Cephalosporin C.

• 한국미생물·생명공학회지
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• 제17권1호
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• pp.46-50
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• 1989

최소 생산배지에 포함된 25mM 이상의 무기인(Inorganic phosphate)은 Cephalosporium acremonium의 생육은 증대시켰으나, cephalosporin C(CPC) 생산은 감소시켰다. Phosphate-starved 휴지세포계를 이용하여 균체내의 ATP, ADP와 AMP를 측정하였다. 그 결과로부터 무기인은 adenylated nucleotide의 농도의 비율로 표시되는 energy charge를 조절하여 CPC 생산성에 영향을 주는 것으로 밝혀졌다. 또한, 무기인에 의한 CPC 생합성은 repression 효과에 의한 것이 아니라 inhibition 효과에 의해 조절받는 것을 알 수 있었다.

• 한국식품영양학회지
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• 제12권3호
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• pp.271-278
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• 1999

We isolated activnmycetes LAM-98-80 as strain producing an effective antibiotic for cephalosporin re-sistant pathogenic PSeudomonas sp. and identified as Streptomyces sp. LAM-98-80 from cultural and phyisological characteristics. We investigated the optimal culture conditions for producation of an anti-biotic becoming effective for cephalsporin-resistant pathogenic Pseudomonas sp. It was found that 1.5% soluble starch and 1.0% yeast extract were good as carbon and nitrogen source respectively. The pro-duction of antibiotic was also activated by 0.04% Mn2+ as 80% degree. The optimum initial pH on pro-ductio of antibiotic was pH 7.0. The culture condition for the maximal productivity of the antibiotic was at 3$0^{\circ}C$ for 5 days. The cephalosporin-resistant pathogenic Pseudomonas sp. as test bacteria was rev-ealed to resist antibiotic of cepha families but revealed to not resist those of $\beta$-lactam families ampicil-lin and amoxicillin. Parital purified antibiotic was stable for the pH from 3 to 9 and was also stable when treated at 70 $^{\circ}C$ for 1 hour, This antbiotic was effective against all gram positive and negative bac-teria but was not effective against molds and yeasts.

• Archives of Pharmacal Research
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• 제22권4호
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• pp.391-397
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• 1999

The quaternary ammonium cephalosporin derivatives were prepared with various pyridines substituted at the 3 or/and 4 position. Their in vitro antibacterial activities were determined and substituent effect on pyridine nucleus was studied. Preparation of substituted pyridines are also described.

• Bulletin of the Korean Chemical Society
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• 제12권6호
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• pp.674-678
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• 1991

Methanolysis of a ${\beta}$-lactam ring of a cephalosporin was simulated with AM1 semiempirical quantum mechanical calculation. The tetrahedral intermediate TD1 from an O-protonated cephalosporin and a methanol transfers the proton intramolecularly to the C-4 carboxylate to generate an oxyanion, i.e., second tetrahedral intermediate TD2, which undergoes the amide bond cleavage without further protonation on the N-5. For this cleavage a low-energy barrier TS2 was located. According to the energy diagram, tetrahedral intermediates easily undergo ring cleavage even without the protonation on the amide nitrogen.

• Journal of Microbiology and Biotechnology
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• 제5권4호
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• pp.229-233
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• 1995

Acremonium chrysogenum was immobilized in ionotropic gel beads to develop semi-continuous production of cephalosporin C (CPC). Barium alginate beads were more stable than calcium alginate or strontium alginate beads in chemically defined media. The gel stability of Ba-alginate was further increased by cross-linking with polyethyleneimine (PEI). The presence of carboxymethyl cellulose inside Ba-alginate beads did not reduce mass transfer resistance. Ba-alginate microbeads that had little diffusion limitation increased CPC production rate 1.6 fold higher than that of normal beads. CPC fermentation with immobilized cells in Ba-alginate microbeads was performed continuously for 40 days by way of repeated fed-batch operations. Mathematical modeling was developed to describe the repeated fed-batch fermentation system. Results of the computer simulation agreed well with the experimental data, which made it possible to predict an optimal feeding rate that could maximize total CPC productions.