tudiea~qn,~pto~lucin ells of p-`futant posscssin d-glucosq tso*meras

8/14/2019 Tudiea~Qn,~Pto~Lucin Ells of P-`Futant Posscssin D-GlucosQ Tso*Meras

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tudiea~qn,~Pto~lucin ells of P-`futant Posscssin D-GlucosQ Tso*meras eACtiVi ty ,'H;,

in the MRR for February, efforts.were being made to fermentan~finexpAnsive,medium .to,obtainArthrohactercells:wi,th high glucoseisomerase. .,_ .. M . . .activi.ty.,;Investigations by Dr . Lee showed that cells of a double-mutant(F-mutant) :'whichhe had obtained possessed high enzymatic activity when culture din&'a`-'rnedi,um 'with'2X dextroseas the c3rbori source : Therefore, production of6011Sof F-mutant was examined .

An inaeulum of F-mutant was prepared by either of two methods. Themutantwas transferred by streaking from"a~stock culture to plates of nutrient aga r

0.6% O .M : HAP,'0 .1% BFY 100, 0.6% (%'H4)2HP04', 0.2% KH2P04 ; 0.0125% MgSO471120 ,

inoculum A: To'prepare inoculum B, the mutantwas cultured for 24 hours (30C) onnutrientagar, and the growth`(1/2 standardloop) was transferred to a medium.(100 in1 at ;;pHk.7 .l5,before autoclyving/500-m1_1flasks):that contained 2% dextrose ,

(Aifco) :, ;,,The;plates:were incubated at 30C for 24 hours . Sterile water (10 ml)was added,;to~-oneplate and with gentle agitation, a cell suspension as prepared .Production.media: were .then inoculated by'a`0.5% transfer of the cell suspension ,

. and_ ;distilled,water:, After24-27 hours on the shaker (30C), produciton medi a

cell'production contained 2% dextrose, 0.6%, 0.r1 .llAP, 0.1 - 0175% BYF 100,0 .6% (N114)2HP04, 0 .2% K112P04, 0 .01 - 0 .025% 1-1gSO471120, and distilled water .The flasks were incubated at 30C on the shaker rotating at 250 rpm with a2-inch stroke .

The media (100 ml at pH 7.15 before autoclaving 500-m1 flask) used fo rtiere"inoculated by a0.5 - 1Z transfer from the shake .

Assays of the cell for glucose isomerase activity showed activities ofas much as 470 - 630 u units/ml after 64 to 112 hours of incubation (Table I) .The yields were considered good but not as high as expected .

In co-operation with Dr . Lee, two 30-gallon fermentations (Nos. 46 and 47)were conducted . Data on those will be reported by Dr . Lee .

One 30-gallon fermentation (No .48) with the 2% dextrose medium as used inshakes was inoculated with 1% of B-type inoculum . The propeller was rotatedat 300 rpm and an aeration rate of 0 .45 cu. feet/r:inute/30 gallons was e;,p]oyed .The results showed a naximum yield of 480 p units/ml after 64 licurs of incubation .The yield (440) was slightly lower at 67 hours . The final p1i w: s 5.4 -Ind oncentrifrigation, 3333 bms of cells were obtained .

Presently, methods to consistently produce cells of F-mutant with highenzymatic activity (7001) are being studied.


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:~~Asslbtud Dr . W . C . Squires . i n ' ' p r o p a r i ' n g ' . t h r c i o , ; ` r e p o : , t s . ' T w o ; ;Bio-

syntheFis of Compound 350 and Treatnent of A ir'-Conditioning Sjstcirls ,' . w o ' r e

R~It rcports and one, Proparation of 1 .ectono of`3-Sucrinoylpyridonc-6, was

an KUM .

P i r . ' :-MAtthews ass isted,Tlise Long with,foo4 ;stud ies and antibiotic studies ; ; 'ral:~tn~l tn tha F..mntnnt_ . , .


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0 .02 50 .6 : '0 .125 551 6260 .6 0 .1 0.6 0.2 0 .0125 B 395 34 5

0 .6 0 .125 0 .6 0 .2 0 .0125 A 421 2040 .6 0 .175 0 .6 0 .2 0.0125 A 432 25 50.6 0 .1 0 .6 0.2 0 .01 A 418 1550.6 0 .1 0 .6 0 .2 0.01 A 421 17 20.6 0 .1 0 .6 0 .2 0 .0125 A 399 2530 .6 0 .1 0 .6 0 .2 0 .0125 A 412 353

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0 .6 0 .2 0.01250:6 0.2 0 .0125 .

6 0 .2 -0 .0125 ;.6 : . 0 .2 0 .0125 : 397: 6=. 2 0. 025 _, . . ' .


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*Incculum A culturod' o n ;:nutrient agar plate for 24 hours at 30C . added

10 ml ateril o . : ;distilicd`wAter'.~ ,Transferred Q,. S m l ` c e l l : cuspansiQnto - :

productionmedia, 100 m1/500 ml" f1ASk .f' Inc+culum`$ ; p C u .itured' on tiutrient

agar plato ; ;for 24 hours at 30'C . .transfcrred :;grow th'(1/2 stAnc~Ard loop) tomedium containing 2Xidaxtroso, 100,:0 . 6 x ~ ' , ( N H ~ ) 2 1 i P 0 4 ,0,2X hH2P0y'0125Y ;1~1gSO4dlstilled'h~aker .+j Incubated on'shakerfor 24 houra,-; _and then inoculated media'by 0, .5 on ;1%''tra nsfer .

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* * 2 X , d e x t` r o s e ; c o n t e nt ~ ` w ~ a s o b t a il l e d ;by a dding 51 .8% dextrose syrup that w,a s

Lawrence E . Hayes ~'

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tudiea~qn,~pto~lucin ells of p-`futant posscssin d-glucosq tso*meras

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