Production and characterization of

thermostable α-amylase from a newly

isolated strain of Bacillus subtilis
KIBGE-HAR
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Aliya Riaz
Pharmaceutical Research Centre
PCSIR Laboratories complex

Shah Ali Ul Qadar

Institute of Sustainable Halophytes Utilization
University of Karachi

Abida Anwar
Pharmaceutical Research Centre
PCSIR Laboratories complex

Samina Iqbal
Pharmaceutical Research Centre
PCSIR Laboratories complex

Saeeda Bano
Pharmaceutical Research Centre
PCSIR Laboratories complex

Citation: A. Riaz, S. Qadar, A. Anwar, S. Iqbal & S. Bano : Production and characterization of
thermostable α-amylase from a newly isolated strain of Bacillus subtilis KIBGE-HAR . The Internet
Journal of Microbiology. 2009 Volume 6 Number 1

Keywords: Fermentation | thermostable | medium | optimization | alpha amylase

Table of Contents
 Introduction
 Materials And Methods
 Organism
 Media Composition
 Production of α -Amylase
 Enzyme Assay and Protein Determi...
 Parametric Analysis for α-amyla...
 Comparison of different media co...
 Characterization of α-amylase
 Results
 Time Course of Cellular Growth a...
 Effect of Starch Concentration o...
 Effect of Temperature on α-amyl...
 Effect of pH on α-amylase produ...
 Selection of buffer for α-amyla...
 Effect of Peptone Concentration ...
 Effect of Yeast Concentration on...
 Effect of Ca+2 on α -amylase Pr...
 Comparison of different media fo...
 Discussion
 Conclusion
 Acknowledgment
 Corresponding Author

Abstract
Investigation on the fermentation conditions for Alpha-amylase (1,4-α-D-glucan
glucanohydrolase, E.C. 3.2.1.1) production was carried out with Bacillus subtilis
KIBGE-HAR and a optimal synthetic medium for enzyme production was developed.
Alpha-amylase production and cell population reached maximum after 24 hours of
cultivation. The optimum temperature and pH for enzyme production were found to be
50°C and 7.0 respectively. Starch (15 g/l), which was used as a carbon source, supported
the maximum production of enzyme. Peptone was used as a nitrogen source and the best
concentration of peptone for α-amylase formation was found to be 5 g/l. High α-amylase
titre was obtained in medium supplemented with 1.0 g/l yeast extract. Calcium chloride
was added in the medium as a stabilizer and 0.2 mg/dl CaCl2 was found to be the most
-amylase production and stability. Among differentfavorable concentration for
-amylase production, medium 3 gave the maximum yieldreference media tested for
-amylaseof enzyme and our own optimized medium 4 was proved to be optimal for
production in contrast with the reference media. The optimum temperature and pH of the
enzyme were found to be 60oC and 7.0 respectively. The most suitable buffer system for
pH maintenance was proved to be Tris-HCl buffer (50 mM). Velocity of reaction
reached to maximum in the presence of 2 % substrate i.e. starch.

Gene Bank Accession: EU819144 (16S rRNA gene sequencing for Bacillus subtilis
KIBGE- HAR)

Introduction
The α-amylase (E.C. 3.2.1.1) randomly hydrolyzes alpha 1,4 glucosidic linkages in
starch, glycogen and related polysaccharides yielding dextrins, oligosaccharides,
maltose and D-glucose (Takeshita et al., 1975). Bacterial α-amylases are extensively
important in industrial processes such as production of ethanol and high fructose corn
syrup, baking, in laundry washing powders and dish washing detergents, textile
desizing, and paper recycling (Nigam and Singh, 1995).

Thermostable enzymes are more versatile rather than thermolabile (Fogarth et al., 1974)
as they have higher operational stability and a longer shelf life at elevated temperatures
(Niehaus et al., 1999). Therefore the thermophilic microorganisms are of special interest
for producing thermostable α- amylase, which can be use in a wide array of industrial
processes (Chandra et al., 1980; McMohan et al., 1997). Bacteria belonging to the genus
Bacillus have been widely used for the commercial production of thermostable α-
amylase. These include B. coagulans, B. stearothermophilus, B. caldolyticus, B. brevis,
B. acidocaldarius and B. thermoamyloliquefaciens (Campbell, 1954, 1955).

Due to the increasing demand for thermostable α-amylase in various industries, it has
been produced and characterized from different sources. The characteristics of α-
amylase, such as its thermostability and pH profile should match its application.

In this regard it is essential to work on the conditions that lead to the bulk production of
thermostable amylase for industrial applications and to search for α-amylase with
improved properties. Therefore, the present study was carried out to optimize the
fermentation conditions for the production of α-amylase from Bacillus subtilis KIBGE-
HAR and to characterize the enzyme produced in the culture supernatant as well as
comparison of different reference media with that of our own optimized medium for the
production of α-amylase.
Materials And Methods
Organism
The strain was isolated from air and pure culture study was performed. A pure culture of
Bacillus subtilis KIBGE-HAR was used and stock culture was maintained on nutrient
agar slant and for alpha amylase production the strain was weekly revived in 5 ml starch
broth.

Media Composition
For α-amylase production, a culture medium was prepared containing (g/l): 15.0 soluble
starch, 1.0 Yeast extract, 5.0 Bacto Peptone, 0.5 MgSO4, 0.5 NaCl and 0.002 CaCl2. The
pH of the medium was adjusted to 7.0 before sterilization (Aliya et al. 2007).

Production of α -Amylase
Starch broth (45 ml) was inoculated with 5 ml of an overnight culture of B. subtilis
KIBGE-HAR and then incubated at 50 °C for 24 hours. After 24 hours, fermented broth
was centrifuged at 10,000 r.p.m for 10 minutes at 0°C. The cell free filtrate (C.F.F.) was
used for enzyme assay.

Enzyme Assay and Protein Determination

The saccharolytic activity of α-amylase in the culture filtrate was assayed by incubating
0.1 ml CFF with 1 ml soluble starch (2 % w/v, prepared in 50 mM Tris-HCl buffer of
pH 7.0) at 60±C for 5 minutes. The α-amylase level was determined by measuring the
reducing sugar released from soluble starch (Nelson, 1944; Somogyi, 1945; Somogyi,
1952).

“An enzyme unit is defined as the amount of α-amylase that liberates 1 µmol of
reducing sugar from the substrate in one minute at 60±C”.

Total Protein was measured by the Lowry et al. method (1951). Bovine serum albumin
(250 µg/ml) was used as a standard.

Parametric Analysis for α-amylase production

The Bacillus subtilis KIBGE-HAR was grown in the medium for 6, 18, 24, 48, 72, and
96 hours for the time course of maximum cellular growth and α-amylase production.
Wet cell mass (g/dl) was measured after each interval of time.

The carbon source used was starch and different concentrations of starch (5 – 30 g/l)
were tested to get the best one for α-amylase production.
The effects of temperature and pH of the medium were also studied. This was carried
out by growing the organism at different temperatures (37 °C – 60 °C) and different pH
values (5 – 10).

Various concentrations of nitrogen source i.e. peptone (0 – 20 g/l) were analyzed for the
maximum production of α-amylase.

The effects of different levels of Yeast extract (0 – 5 g/l) and CaCl2 (0.001 – 0.01 g/l)
were also investigated.

Comparison of different media compositions for α-

amylase production from Bacillus subtilis KIBGE-
HAR
Four different fermentation media were used for α-amylase production from Bacillus
subtilis KIBGE-HAR in order to compare the level of α-amylase produced in them with
that of our own optimized medium (Table 1).

Table 1: Media compositions for α-amylase production from Bacillus subtilis KIBGE-
HAR

(Initial pH values of the media were adjusted to 7.0 before sterilization)

Characterization of α-amylase
Effect of substrate concentration on enzyme activity was measured at different
concentrations of starch in the reaction mixture (0.25% - 2.5 %)

The temperature optimum was evaluated by performing the enzyme assay at different
temperatures ranging from 35 °C to 75 °C.

Effect of pH on the activity of α-amylase was determined by measuring the activity at

different pH values ranging from pH 5.5 to 9.0.

For the selection of appropriate buffer to get maximum enzyme activity, different buffer
systems (50 mM each) including Citrate-Phosphate buffer, Phosphate buffer and Tris-
HCl buffer of pH 7.00 were used.
Results
Time Course of Cellular Growth and α-amylase
Production
A linear relationship was found between enzyme synthesis and cell growth i.e.
maximum α-amylase production occurred when the cell mass reached to maximum (Fig.
1)

Figure 1: Time course for α-amylase production and wet cell mass from Bacillus subtilis
KIBGE-HAR

Effect of Starch Concentration on α-amylase

Production and extracellular activity
Figure-2 showed that medium containing 1.5 g% starch supported the maximum
production of α-amylase and increasing starch concentration beyond 1.5 g% resulted in
the declined enzyme production.

Figure 2: Effect of starch concentration on α-amylase production from Bacillus subtilis

KIBGE-HAR

Figure-3 showed the graphical analysis of the effect of substrate concentration on

extracellular α-amylase activity. The maximum velocity was achieved with 2 % starch,
where after it declined sharply.
Figure 3: Effect of substrate concentration on extracellular α-amylase activity

Effect of Temperature on α-amylase production and

extracellular activity
It was found that enzyme production in the fermentation medium increased with
increase in temperature and maximum enzyme production was obtained at 50°C and
after, enzyme production was sharply decreased at 55°C. At 50°C the maximum
bacterial cells multiplication occurred and during their multiplication they secreted
extracellular enzyme (Fig. 4).

Figure 4: Effect of temperature on α-amylase production during fermentation from

Bacillus subtilis KIBGE-HAR

The optimum temperature for maximum extracellular α-amylase activity was found to
be 60°C.

Further increase in temperature resulted in reduce enzyme activity (Fig. 5).

Figure 5: Effect of temperature on extracellular α-amylase activity

Effect of pH on α-amylase production and extra

cellular activity
Maximum enzyme production was observed when pH of the medium was kept 7.0
before sterilization (Fig. 6), while at pH 5 and pH 9, 68 % and 30 % production was
observed respectively, with reference to the optimum pH.

Figure 6: Effect of initial medium pH on the production of α-amylase from Bacillus

subtilis KIBGE-HAR

The effect of pH on α-amylase activity is shown in Fig-7. Enzyme showed maximum

extracellular α-amylase activity at neutral pH and as the pH increases the enzyme lost its
activity.

Figure 7: Effect of pH on the extracellular α-amylase activity

Selection of buffer for α-amylase activity

Among different buffers tested, Tris-HCl buffer (50 mM) provided the most suitable
buffering environment in which α-amylase activity was maximum (Fig. 8).

Figure 8: Effect of different buffers (pH 7.00) on extracellular α-amylase activity

Effect of Peptone Concentration on α-amylase

Production
The effect of peptone on α-amylase production by Bacillus subtilis KIBGE-HAR
showed that enzyme production was effected by the change in concentration of peptone
in the medium (Fig. 9). Maximum α-amylase production was observed when 0.5 g%
peptone was added in the medium.
Figure 9: Effect of Peptone concentration in medium on α-amylase production from
Bacillus subtilis KIBGE-HAR

Effect of Yeast Concentration on α-amylase Production

It was observed that medium containing 0.1 g% yeast extract supported optimal α-
amylase production (Fig. 10) followed by a decline at high concentrations.

Figure 10: Effect of yeast concentration in medium on α-amylase production from

Bacillus subtilis KIBGE-HAR

Effect of Ca+2 on α -amylase Production

Figure 11 showed that when 0.2 mg/dl CaCl2 was added in the fermentation medium,
resulted in the enhancement of α- amylase production.

Figure 11: Effect of CaCl2 concentration in medium on α-amylase production from

Bacillus subtilis KIBGE-HAR

Comparison of different media for α-amylase

production from B. subtilis KIBGE-HAR
It has been observed that the strain of B.subtilis KIBGE-HAR produced 631 U/ml/min in
medium-3 while 844 U/ml/min in medium-4, which was our own synthetic medium
developed for optimal enzyme production. Medium-2 showed the least α-amylase yield
as compare to others.
Discussion
Cellular growth of B. subtilis KIBGE-HAR and α-amylase production was affected by
the time of incubation. Growth dependent α-amylase production was found by this strain
i.e., a linear relationship was found between enzyme synthesis and cell growth. It was
observed that maximum α-amylase production occurred when the cell mass reached to
its maximum. This suggests that this organism may be sensitive to metabolite repression
(Cordeiro et al., 2002). It was also known that optimum temperature for growth and α-
amylase production was same. Other investigators also reported the same as above
(Bajpai and Bajpai, 1989; Saito and Yamamoto, 1975).

Different carbon sources affect differently on the production of α-amylase (Welker and
Campbell, 1963) and the most commonly used carbon source is starch (Bajpai and
Bajpai, 1989). Starch at concentration of 15 g/l supported maximum production of
enzyme followed by a decline at high concentrations. This may be the effect of catabolic
repression i.e. glucose which was formed during the hydrolysis of starch may influenced
negatively on α-amylase gene expression. These results are similar to the findings of
Haseltine et al. (1996) who observed that glucose repressed the production of amylase
by the hyperthermophilic Archaeon Sulfolobus solfataricus.

Starch was also used as a substrate for α-amylase activity. Optimum activity was
achieved at 2 % starch, and as the substrate concentration increased, α-amylase activity
dropped sharply which may be due to the substrate inhibitory effect.

Fermentation temperature has a profound effect on the level of enzyme produced in the
medium. In the present study, α-amylase production by B. subtilis KIBGE-HAR was
found to be maximum at 50 °C where after it declined. Amylase production by many
Bacillus sp. is known to be affected by the amount of dissolved oxygen in the medium
(Wind et al., 1994). At elevated temperature, the solubility of oxygen is decreased in the
medium (Campbell and Pace, 1968) leading to limitation of dissolved oxygen for the
production of α-amylase, as it has been reported that difference in the level of oxygen in
the medium induced changes in the surface protein layer (S-layer) of bacterial cell
membrane (Sara et al., 1996) which is involved in the control of extracellular release of
enzyme. This may be the reason of decreased amylase production beyond 50 °C.

Temperature is also the most important factor which markedly influences the enzyme
activity. In the present study, optimum temperature for α-amylase activity was 60 °C.
Further increase resulted in reduced enzyme activity as high temperatures induce
conformational changes in three dimensional structure of an enzyme and its
accommodation to substrate molecules becomes lower.

Medium pH also affects α-amylase production. Composition of cell wall and plasma
membrane of microorganisms is known to be affected by the medium pH (Ellwood and
Tempest, 1972). Due to this change in the nature of cell wall and plasma membrane the
growth parameters may vary, especially temperature, and the provided temperature may
not remain suitable for the growth of organism (Stutzenberger and Jenkins, 1995). This
may be the reason of decreased production of α-amylase at pH 5 and 9 in the present
study.

Optimum pH for α-amylase activity was found to be 7.0. The enzyme activity at pH 6.0
and 7.5 were 65 % and 77 % of that at pH 7.0, respectively. At pH 8.0, enzyme activity
reduced to 46 %. Thus, we can conclude that α-amylase from Bacillus subtilis KIBGE-
HAR is active at or near neutrality.

The nature and concentration of nitrogen source are important for the formation of α-
amylase. Lower level of nitrogen is inadequate for the enzyme production and excess
nitrogen is equally detrimental causing enzyme inhibition (Dharani Aiyer P. V., 2004).

As a nitrogen source peptone was added for α-amylase production. Similar finding was
also reported previously (Davies et al, 1980).

The concentration of yeast extract was found to be crucial for the production of α-
amylase and as the concentration increased after reaching maxima in medium a sharp
fall of enzyme production was observed. The reason may be that the high concentration
of yeast decreased the pH of the medium during fermentation which ultimately destroys
the enzyme produced in the medium (Babu and Satyanarayana, 1993). Another reason
could be that both yeast extract and peptone affect the surface charge, hydrophobicity
and nitrogen-to-carbon ratio of the bacterial cell wall (Schar-Zammaretti Prisca et al.,
2005) and the hydrophobicity of the cell wall decreases as the concentration of yeast
extract and peptone increase in the medium. This decreased hydrophobicity of bacterial
cell wall may result in decreased extracellular release of enzyme.

Calcium ions are known to be the stabilizer and activator of α-amylase and it has been
reported that requirement of Ca+2 is different for thermostable α-amylase as compared
to thermolabile. In case of thermolabile α-amylase, Mamo and Gassesse (1999) reported
that 0.1 g/l CaCl2 was optimum for amylase production by Bacillus sp. WN 11 whereas
Qader et al. (2006) reported that in case of Bacillus sp. AS-1, 0.2 g/l CaCl2 was optimum
for the maximum production of α-amylase. In the present study, thermostable strain
required low quantity of 0.002 g/l CaCl2 for optimum production of α-amylase.
Similarly Welker and Campbell (1963) also worked on thermostable α-amylase and used
0.005 g/l CaCl2 in the fermentation medium. This suggests that thermostable α-amylase
has a low requirement for calcium ions as compared to thermolabile or we can say that
thermostable α-amylase possesses high affinity for calcium ions (Kindle Karen et al.,
1986).

Different media have been compared with that of our own optimized medium i.e.
medium 4. Medium 2 reduced markedly the enzyme production which may be due to the
greater amount of yeast extract present and the strain did not utilized this extra yeast
extract in the fermentation medium and due to this high concentration of yeast extract
inhibition started (Babu and Satyanarayana, 1993). Medium 1 contained tryptone as a
nitrogen supplement and showed lower enzyme production as compare to medium 3 and
4 which did not contain tryptone but peptone. It has been reported that maximum
enzyme production was found with peptone as the nitrogen source (Lin et al., 1998,
Dharani Aiyer, 2004) and tryptone resulted in decreased enzyme production. This may
be the reason why medium 1 did not produced α-amylase optimally.
Media 1 and 2 which showed reduced enzyme production as compare to media 3 and 4
and did not contain CaCl2. It was previously reported that Ca+2 is known to be the
stabilizer and activator of α-amylase (Kennedy et al., 1979; Hewitt et al, 1996), therefore
the absence of CaCl2 in media 1 and 2 might be responsible for low enzyme production.

Conclusion
In the present study, we developed the new medium composition for the maximum
production of α-amylase from B. subtilis KIBGE-HAR. Enzyme characterization was
performed which indicates that enzyme is thermostable and showed optimum activity at
60°C. This alpha amylase maintains its activity upto high temperature and can easily be
used for industrial purpose. Enzyme also showed 20% and 40% loss of activity at 65°C
and 70°C respectively.

Acknowledgment
Authors acknowledge the help provided by Dr. Tanzeel Haider Usmani in this study.

Corresponding Author
Dr. SHAH ALI UL QADER
Institute of Sustainable Halophytes Utilization
University of Karachi,
Karachi-75270
Pakistan
Ph # : 92-03212160109
Fax # : 92-021-2229310
E.mail: [madar_chem@yahoo.com]

References
Aliya R., Qader SA, Anwar A, Iqbal S, Bano S (2007) Effect of medium composition
and time course on the production of alpha-amylase from Bacillus stearothermophilus.
Pak. J. Biochem. Mol. Biol. 40(2): 51-54. (s)

Babu KR, Satyanarayana T (1993) Extracellular calcium inhibited α-amylase of Bacillus

coagulans B49. Enzyme Microbial Technol. 15: 1066-1069 (s)

Bajpai P, Bajpai PK (1989) High-temperature alkaline α-amylase from Bacillus

licheniformis TCRDC-B13. Biotech. Bioeng. 33(1): 72-78 (s)

Campbell LL (1954) Crystallization of α-amylase from a thermophilic bacterium. J.

Amer Chem Soc. 76: 52-56 (s)

Campbell LL (1955) Purification and properties of α-amylase from facultative

thermophilic bacteria. Archives of Biochemistry and Biophysics. 54: 24-35 (s)

Campbell LL, Pace B (1968) Physiology of growth at high temperature. Journal of Appl.
Bacteriology 31: 24-35 (s)

Chandra AK, Medda S, Bhadra AK (1980) Production of extracellular thermostable α-

amylase by Bacillus licheniformis. J. Ferment. Technol. 58: 1–10 (s)

Cordeiro CAM, Martins MLL, Luciano AB (2002) Production and properties of α-

amylase from thermophilic Bacillus sp. Braz. J. Microbiol. 33: 57-61 (s)

Davies PE, Cohen DL, Whitaker A (1980) The production of α-amylase in batch and
chemostat cultures of Bacillus stearothermophilus . Antonie Van Leeuwenhoek 46 (4):
391- 398 (s)

Dharani Aiyer PV (2004) Effect of C:N ratio on α-amylase production by Bacillus

licheniformis SPT 27 African Journal of Biotechnology. 3(10): 519-522 (s)

Ellwood DC, Tempest DW (1972) Effects of environment on bacterial wall content and
composition. Advances in Microbial Physiology 7: 83-117 (s)

Fogarth WM, Griffin PJ, Joyce AM (1974) Enzyme of Bacillus species – part 1. Process
Biochem 9: 11-24 (s)

Haseltine C, Rolfsmeier M, Blum P (1996) The glucose effect and regulation of α-

amylase synthesis in the hyperthermophilic archaeon Sulfolobus solfataricus. J.
Bacteriol. 178: 945-950 (s)

Hewitt CJ, Solomons GL (1996) The production of α-amylase ( E.C. 3.2.1.1) by Bacillus
amyloliquefaciens, in a complex and a totally defined synthetic medium . J. Ind.
Microbiol. 17: 96-99. (s)

Kennedy JF, White CA (1979) Stability and kinetic properties of magnetic immobilized
α-amylase. Starch/Staerke. 31: 375-381 (s)

Kindle KL, Mainzer SE, Marlatt DL, Sawyer CB (1986) Thermostable alpha amylase
having a low requirement for calcium ions, derived from a bacillus microorganism.
United States Patent 4600693 (s)

Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the
Folin phenol reagent. J. Biol. Chem. 193: 265-275 (s)

Mamo G, Gessesse A (1999) Effect of cultivation conditions on growth and α-amylase

production by a thermophilic Bacillus sp. Letter in Applied Microbiology 29: 61-65 (s)

McMahon HEM, Kelly CT, Fogarty WM (1999) Thermostability of three α-amylases of

Streptomyces sp. IMD 2679. J. Ind. Microbiol. Biotechnol. 22: 96-99 (s)

Nelson N (1944) A photometric adaptation of Somogyi method for the determination of

glucose. Journal of Biological Chemistry. 153 : 375-380 (s)

Niehaus F, Bertoldo C, Kahler M, Antranikian G (1999) Extremophiles as a source of

novel enzymes for industrial application. Appl. Microbiol. Biotechnol. 51(6): 711-729
(s)

Nigam P, Singh D (1995) Enzyme and microbial system involved in starch processing.
Enzyme and Microb. Technol. 17(9): 770-778 (s)

Qader SA, Bano S, Aman A, Syed N, Azhar A (2006) Enhanced production and
extracellular activity of commercially important amylolytic enzyme by a newly isolated
strain of Bacillus sp. AS-1. Turk J. Biochem. 31(3): 135-140 (s)

Ramachandran S, Patel AK, Nampoothiri KM, Chandran S, Szakacs G, Soccol CR,

Pandey A (2004) α-amylase from a fungal culture grown on oil cakes and its properties.
Braz. Arch. Biol. Technol. 47: 309-317 (s)

Saito N, Yamamoto K (1975) Regulatory factors affecting α-amylase production in

Bacillus licheniformis. J. Bacteriol. 121(3): 848-856 (s)

Sara M, Kuen B, Mayer HF, Mandl F, Schuster KC, Sleytr UB (1996) Dynamics in
oxygen-induced changes in S-layer protein synthesis from Bacillus stearothermophilus
PV72 and the S-layer-deficient variant T5 in continuous culture and studies of the cell
wall composition. Journal of Bacteriology. 178(7): 2108-2117 (s)

Schar-Zammaretti P, Dillmann ML, D'Amico N, Affolter M, Ubbink J (2005) Influence

of fermentation medium composition on physicochemical surface properties of
Lactoibacillus acidophilus , Appl. Envior. Microbiol. 71: 8165-8173 (s)

Somogyi M (1945) A new reagent for the determination of sugar. J. Biol. Chem. 160,
61-68 (s)

Somogyi M (1952 ) Notes on sugar determination. J. Biol. Chem. 195, 19-23 (s)

Stutzenberger FJ, Jenkins TC (1995) Effect of carbon source on growth, temperature and
fatty-acid composition in Thermomonospora curvata. World Journal of Microbiol.
Biotechnol. 11(6): 621-624 (s)

Takeshita M, Hehre E (1975) The capacity of Alpha-amylase to catalyze the

nonhydrolytic degradation of starch and glycogen with formation of novel glycosylation
products, Arch Biochem. Biophys. 169: 627-637 (s)

Welker NE, Campbell LL (1963) Effect of carbon sources on formation of α-amylase by

Bacillus stearothermophilus. J. Bacteriol. 86 (4): 681-686 (s)

de vrij W, Speelmans G, Heyne RIR, Konings WN (1990) Energy transduction and

amino acid transport in thermophilic aerobic and fermentative bacteria. FEMS
Microbiology Reviews 75(2-3) 183-200. (s)
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