Öz
Bu çalışma, sıçan karaciğerinden özütlenen alkalen fosfataz (ALP) enziminin kısmen özütlenmesini, inhibitörsüz ve inhibitör varlığında kinetik özelliklerinin araştırılmasını kapsamaktadır. Deneysel sonuçlar yardımıyla Michaelis Menten ve Lineweaver Burk grafikleri çizildi. Bu grafiklerden Km ve Vmax değerleri hesaplandı. Değerler: İnhibitörsüz Km= 0,047 Vmax=24,87; Üre inhibitörlü Km= 0,054 Vmax=20,83 ; Kreatinin inhibitörlü Km= 0,037 Vmax=20,83 olarak hesaplanmıştır. Bu değerlere göre enzimin substrata ilgisi ve maksimum hız tespit edildi. İnhibitör maddeyle deneyler tekrarlandı ve inhibisyon tipi belirlendi. Kandaki yüksek üre ve kreatinin seviyesinin alkalen fosfataz enzimini nasıl etkilediği bulundu. Kandaki ALP (Alkalen Fosfataz) üzerinde ürenin
inhibitör olarak kullanıldığı çalışmada reaksiyonun yarışmasız (nonkompetetif) bir reaksiyon, kreatininin inhibitör olarak kullanıldığı çalışmada ise reaksiyonun yarışmacı olmayan (unkompetetif) bir reaksiyonla gerçekleştiği tespit edilmiştir.
Anahtar Kelimeler
Alkalen fosfataz Michaelis Menten yarışmasız inhibisyon yarışmacı olmayan inhibisyon.
Kaynakça
- Jakob A., Bryjak J., Wojtowicz H., Illeova V.,
- Annus J., Polakovic M., “Inactivation kinetics
- of food enzymes during ohmic heating”, Food
- Chemistry, No 123, 369–376, 2010.
- Biasutti M., Abuin E., Silber J., Correa M., Lissi
- E., “Kinetics of reactions catalyzed by enzymes
- in solutions of surfactants”, Advances in
- Colloid and Interface Science, No 136, 1–24,
- -
- Wilin´ska A., Bryjak J., Illeova V., Polakovic
- M., “Kinetics of thermal inactivation of alkaline
- phosphatasein bovine and caprine milkand
- buffer”, International Dairy Journal, No 17,
- –586, 2007.
- Ásgeirsson B., Guðjónsdóttir K., “Reversible
- inactivation of alkaline phosphatase from
- Atlantic cod (Gadus morhua) in urea”,
- Biochimica et Biophysica Acta, No 1764, 190–
- , 2006.
- Van Hoof V., De Broe M.E., “Interpretation and
- Clinical significance of alkaline phosphatase
- isoenzyme patterns.” Critical Reviews in
- Clinical Laboratory Sciences., 197-293, 1994.
- B Behnke, M Kemper, H Kruse and D Muller-
- Wiefel, “Bone alkaline phosphatase in children
- with chronic renal failure”, Nephrology
- Dialysis Transplantation, No 13, Oxford
- University Press, Yayın 3, 662-667, 1998.
- Becq F., Fanjul M., Merten M., “Possible
- regulation of CFTR-chloride channels by
- membrane-bound phosphates in pancreatic duct cells”, Proceeding of the National Academys
- of Sciences of the United States of America
- (PNAS), No 327, 337-342, 1993.
- Day A. P., Feher M. D., Chopra R.,
- “Triglyceride fatty acid chain lenght influences
- the post prandial rise in serum intestinal alkaline
- phosphatase activity”, Annals of Clinical
- Biochemistry, No 29, 287-291, 1992.
- Garrett RH, Grisham CM., Biochemistry,
- Second Edition, Saunders College Publishing,
- -427,1999.
- Lehninger, A.L., Nelson, D.L., Cox, M.M.,
- Lehninger principles of biochemistry, New
- York: W.H. Freeman, 2005.
- Leskovac, V., Comprehensive enzyme
- kinetics, New York: Kluwer Academic/Plenum
- Pub.,2003.
- Walsh R, Martin E, Darvesh S., “Limitations of
- conventional inhibitor classifications”, Integr
- Biol (Camb)., No 3, 197-201,2011.
- Stephenson FH., Calculations for Molecular
- Biology and Biotechnology, Academic Pres,
- An Imprint of Elsevier, Amsterdam, 2003.
- Sutton R, Rockett B, Swindells P., Chemistry
- for the Life Sciences, 1st ed., London, 2000.
- Burtis CA, Ashwood ER, Bruns DE, Tietz
- textbook of clinical chemistry and molecular
- diagnosis, 4th ed. St. Louis: Elsevier Saunders,
- -
- Alan Wu., Tietz Clinical Guide to Laboratory
- Tests, 4th ed. Elsevier Saunders, 2006.
Öz
This study scopes the purification of alkaline phosphatase enzyme extracted from the liver and the investigation
of kinetic properties for the reactions with and without inhibitor. Michaelis Menten and Lineweaver Burk graphs
were drawn by means of experimental results. Km and Vmax values were calculated from these graphics.
According to these values, substrate affinity (Km) and maximum velocity (Vmax) values were determined. Values
without inhibitor; Km= 0,047 Vmax=24,87; with urea inhibitor Km= 0,054 Vmax=20,83 ; with creatinin inhibitor
Km= 0,037 Vmax=20,83. The experiments were repeated with inhibitory substance and inhibition type was
established. How high blood urea and creatinine levels affected alkaline phosphatase activity were determined.
In this study, urea was found to cause noncompetetive inhibition and creatinine to cause uncompetetive
inhibition on liver ALP.
Anahtar Kelimeler
Alkaline phosphatase Michaelis Menten Lineweaver Burk noncompetetive inhibition uncompetetive inhibition
Kaynakça
- Jakob A., Bryjak J., Wojtowicz H., Illeova V.,
- Annus J., Polakovic M., “Inactivation kinetics
- of food enzymes during ohmic heating”, Food
- Chemistry, No 123, 369–376, 2010.
- Biasutti M., Abuin E., Silber J., Correa M., Lissi
- E., “Kinetics of reactions catalyzed by enzymes
- in solutions of surfactants”, Advances in
- Colloid and Interface Science, No 136, 1–24,
- -
- Wilin´ska A., Bryjak J., Illeova V., Polakovic
- M., “Kinetics of thermal inactivation of alkaline
- phosphatasein bovine and caprine milkand
- buffer”, International Dairy Journal, No 17,
- –586, 2007.
- Ásgeirsson B., Guðjónsdóttir K., “Reversible
- inactivation of alkaline phosphatase from
- Atlantic cod (Gadus morhua) in urea”,
- Biochimica et Biophysica Acta, No 1764, 190–
- , 2006.
- Van Hoof V., De Broe M.E., “Interpretation and
- Clinical significance of alkaline phosphatase
- isoenzyme patterns.” Critical Reviews in
- Clinical Laboratory Sciences., 197-293, 1994.
- B Behnke, M Kemper, H Kruse and D Muller-
- Wiefel, “Bone alkaline phosphatase in children
- with chronic renal failure”, Nephrology
- Dialysis Transplantation, No 13, Oxford
- University Press, Yayın 3, 662-667, 1998.
- Becq F., Fanjul M., Merten M., “Possible
- regulation of CFTR-chloride channels by
- membrane-bound phosphates in pancreatic duct cells”, Proceeding of the National Academys
- of Sciences of the United States of America
- (PNAS), No 327, 337-342, 1993.
- Day A. P., Feher M. D., Chopra R.,
- “Triglyceride fatty acid chain lenght influences
- the post prandial rise in serum intestinal alkaline
- phosphatase activity”, Annals of Clinical
- Biochemistry, No 29, 287-291, 1992.
- Garrett RH, Grisham CM., Biochemistry,
- Second Edition, Saunders College Publishing,
- -427,1999.
- Lehninger, A.L., Nelson, D.L., Cox, M.M.,
- Lehninger principles of biochemistry, New
- York: W.H. Freeman, 2005.
- Leskovac, V., Comprehensive enzyme
- kinetics, New York: Kluwer Academic/Plenum
- Pub.,2003.
- Walsh R, Martin E, Darvesh S., “Limitations of
- conventional inhibitor classifications”, Integr
- Biol (Camb)., No 3, 197-201,2011.
- Stephenson FH., Calculations for Molecular
- Biology and Biotechnology, Academic Pres,
- An Imprint of Elsevier, Amsterdam, 2003.
- Sutton R, Rockett B, Swindells P., Chemistry
- for the Life Sciences, 1st ed., London, 2000.
- Burtis CA, Ashwood ER, Bruns DE, Tietz
- textbook of clinical chemistry and molecular
- diagnosis, 4th ed. St. Louis: Elsevier Saunders,
- -
- Alan Wu., Tietz Clinical Guide to Laboratory
- Tests, 4th ed. Elsevier Saunders, 2006.
Ayrıntılar
| Birincil Dil | Türkçe |
|---|---|
| Konular | Mimarlık |
| Bölüm | Makaleler |
| Yazarlar | |
| Yayımlanma Tarihi | 27 Mart 2014 |
| Gönderilme Tarihi | 27 Mart 2014 |
| Yayımlandığı Sayı | Yıl 2013 Cilt: 28 Sayı: 1 |