Bacillus lentus

Several new detergent enzymes have emerged on the market (Table 1). Truly alkaline proteases, introduced in 1974 and 1982, were fermented on strains of Bacillus lentus firmus. These enzymes have a pH optimum between 9 and 11, and have taken important market shares from Alcalase. 

Methyl-60 ,90fBacterium Bacillus lentus Valeric acid chloride... 

For this purpose a fermenter made of stainless steel having a 50 liter capacity is charged with 30 liters of a nutrient solution of 0.1% yeast extract, 0.5% cornsteep and 0.2% glucose, heated for one-half hour at 120°C for sterilization purposes, and after cooling, inoculated with a bacterial suspension of Bacillus lentus MS 2B4. 

Aspergillus sclerotiorum Huber Oxamniquina Bacillus lentus Dasoximatasona Diflucortolona valarata Bacillus polymyxa Polymyxin... 

Plettner E., DeSantis G., Stabile M. and Jones J. B. (1999) Modulation of esterase and amidase activity of subtilisin Bacillus lentus by chemical modification of cysteine mutants. J. Am. Chem. Soc. 121, 4977-4981. 

Bacterium Curvularia lunata 16a-Methyldesoxycorticosterone Bacterium Bacillus lentus Glucose... 

Bacterium Bacillus lentus Valeric acid chloride... 

A hetero-Diels-Alder reaction has been used to prepare racemic 2-ethoxycarbonyl-3,6-dihydro-2H-pyran (9). This ester 9 was resolved by Bacillus lentus protease to provide the R-isomer. Reduction, protection, and ozonolysis provided the bis-mesylate 10 (Scheme 26.9), a key intermediate in the synthesis of the PKC (protein kinase C) inhibitor LY333531 (ll).278 This resolution approach was used because it was more efficient than an asymmetric Diels-Alder reaction. 

Graycar, T., Knapp, M., Ganshaw, G., Dauberman, J., and Bott, R. (1999). Engineered Bacillus lentus subtilisins having altered flexibility. Mol. Biol., 292(1), 97—109. 

OAc yo rff Aspergillus ochraeeus and Bacillus lentus (mixed culture) 68 203... 

The subtilisins used in today s detergent products are usually from Bacillus lentus and other Bacillus species. The survival and growth of these soil-dwelling organisms involves secreting large amounts of... 

Bacillus coagulans Bacillus lentimorbis Bacillus lentus Bacillus licheniformis Bacillus megaterium Bacillus mycoides Bacillus pasteurii Bacillus pseudomycoides Bacillus subtilis Bacillus thuringiensis Bacillus weithenstephanensis... 

A cost effective and easily scaled-up process has been developed for the synthesis of (S)-3-[2- (methylsulfonyl)oxy ethoxy]-4-(triphenylmethoxy)-1 -butanol methanesulfonate, a key intermediate used in the synthesis of a protein kinase C inhibitor drug through a combination of hetero-Diels-Alder and biocatalytic reactions. The Diels-Alder reaction between ethyl glyoxylate and butadiene was used to make racemic 2-ethoxycarbonyl-3,6-dihydro-2H-pyran. Treatment of the racemic ester with Bacillus lentus protease resulted in the selective hydrolysis of the (R)-enantiomer and yielded (S)-2-ethoxycarbonyl-3,6-dihydro-2H-pyran in excellent optical purity, which was reduced to (S)-3,6-dihydro-2H-pyran-2-yl methanol. Tritylation of this alcohol, followed by reductive ozonolysis and mesylation afforded the product in 10-15% overall yield with excellent optical and chemical purity. Details of the process development work done on each step are given. 

Four additional commercial proteases were examined for the hydrolysis of (R,S)-4 including three variants of Bacillus lentus proteases engineered for stability (Tab. 2). Of these Bacillus lentus protease-I gave equivalent selectivity to that of Ba-... 

The effect of pH on the reaction rate and enantioselectivity using Bacillus lentus protease-III was studied and the results are summarized in Tab. 3. As with most ester resolutions using alkaline proteases, at high pH the hydrolysis is more rapid, however the selectivity is somewhat lower. We also observed that base-catalyzed hydrolysis is significant when the pH is over 9.0 (<1%), which has a negative effect on selectivity. Based on initial rate measurements, the rate of hydrolysis at pH 8 was four-fold higher than that at pH 7. Thus there is a substantial decrease in rate of hydrolysis with decreasing pH. 

Reaction conditions room temperature, 1.0 g substrate, 8.7 mL 0.3 M phosphate buffer, 0.3 ml Bacillus lentus protease-III. 

The results presented above clearly indicate that the resolution of (R,S)-4 can be easily carried out using Bacillus lentus proteases. As has been mentioned before we chose Bacillus lentus protease-III for further development because of its efficacy and lower cost. Commercial formulations of this enzyme are stable and avail-... 

To a 20 mL flask was added 2.5 g of (R,S)-2-ethoxycarbonyl-3,6-dihydro-2H-pyran, followed by 7 mL of 0.2 M pH 7.5 phosphate buffer and 2 mL of Bacillus lentus protease-III solution (approximately 5% solution of the protein). The biphasic solution was stirred at room temperature (23 °C) using a magnetic stirrer. The pH was checked at 0.5 hour intervals and readjusted to 7.5 by the drop-wise addition of 1 N NaOH (approximately 7 mL were required over the complete reaction). The progress of the reaction was monitored by chiral gas chromatography. After 5 hours, the enantiomeric purity of the unreacted ester was >99% and the reaction was stopped by the addition of 10 mL of MTBE. The pH of the aqueous phase was adjusted to 8.5 and the mixture was transferred into a separatory funnel. The aqueous phase was extracted twice with 20 mL of MTBE and the combined organic layers were extracted once with saturated sodium bicarbonate (10 mL), followed by saturated sodium chloride solution (10 mL), and the organic... 

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