Karpuz (citrullus vulgaris) alfa-galaktozidazının doğal ve sentetik polimerlerde immobilizasyonu
Küçük Resim Yok
Tarih
2000
Yazarlar
Dergi Başlığı
Dergi ISSN
Cilt Başlığı
Yayıncı
Ege Üniversitesi
Erişim Hakkı
info:eu-repo/semantics/openAccess
Özet
V ÖZET KARPUZ (CITRULLUS VULGARIS) a-GALAKTOZİDAZININ DOĞAL VE SENTETİK POLİMERLERDE İMMOBİLİZASYONU TATAR ÖNAL, Seçil Doktora Tezi, Biyokimya Bölümü Tez Yöneticisi: Prof. Dr. Azmi TELEFONCU Ekim 2000, 190 sayfa a-Galaktozidazlar (a-D-galaktozid galaktohidrolaz, EC 3.2.1.22), doğada oldukça yaygın olarak bulunan hidrolaz sınıfi enzimlerdir. Bitkisel, hayvansal ve mikrobiyal kaynaklardan çok çeşitli tekniklerle izole edilerek saflaştınlan bu enzimler, galaktooligosakkaritler ve polisakkaritlerde bulunan a-l,6-galaktoz birimlerini hidrolizlerler. özellikle şeker endüstrisinde önemli bir potansiyele sahip olan ot-galaktozidazlar, kompleks karbohidratlann biyolojik fonksiyonlarının belirlenmesinde, organik sentezlerde, yapı analizlerinde ve medikal alanda oldukça önemli uygulama alanları olan bir enzimdir. Bu nedenle çeşitli kaynaklardan enzimin ekonomik ve verimli bir şekilde izolasyonu, saflaştırılması ve karakterizasyonu son yıllarda üzerinde en çok durulan konular arasındadır. Tüm bunlara rağmen enzim ile yapılan immobilizasyon çalışmaları ise yok denecek kadar azdır. Bu çalışmada, karpuzdan (Citrullus vulgaris) kısmi olarak saflaştınlan a-galaktozidazın doğal ve sentetik polimerlerde kovalent bağlama ile immobilizasyonu gerçekleştirilmiştir. Karpuz, kolay temin edilebilmesi, ekonomik bir kaynak olması ve enzim aktivitesi açısındanzengin olma» nedeniyle tercih edilmiştir. Karpuz a-galaktozidazı, doğal taşıyıcılardan kitin ve sentetik taşıyıcılardan iyon değiştirici reçinede (AmberHte IRA-938) immobilize edilmiştir. Kullanılan kovalent bağlama ile immobilizasyon metodlan ve taşıyıcı tipleri kıyaslanarak, belirlenen uygun metodla immobilize edilen a-galaktozidazm fiziksel ve kimyasal karakterizasyonu gerçekleştirilmiştir. Taşıyıcılar önce l,r-karbonildiimidazol (CDI) ile aktive edilmiş, ardından enzim ya direkt olarak yada uzatıcı bir kol (y-ammobütirikasit; GABA) ile taşıyıcıda immobilize edilmiştir. İmmobilizasyon çalışmalarının optimizasyonu yapıldıktan sonra elde edilen sonuçlar doğrultusunda, CDI ile aktive edilen taşıyıcılardan kitinde direkt olarak immobilize edilen a-galaktozidaz enziminin fiziksel ve kimyasal karakterizasyonu gerçekleştirilmiştir. Enzim aktivitesine bazı parametrelerin (protein miktarı, inkübasyon süresi, pH, sıcaklık ve substrat konsantrasyonu) etkisi incelenerek kararlılık testleri (termal, pH, depo ve operasyonel kararlılık) yapılmıştır. Bunların yanısıra immobilize enzimin tekrar kullanılabilirliği test edilerek rafinoz hidrolizinde kullanılabilecek uygun reaktör tipi belirlenmiştir. Anahtar sözcükler Amberlite IRA-938, Citrullus vulgaris, a-galaktozidaz, immobilizasyon, iyon değiştirici reçine, karpuz, kitin, protein izolasyon ve saflaştırılması, rafinoz hidrolizi
VII ABSTRACT IMMOBILIZATION OF WATERMELON (C1TRULLUS VULGARIS) a-GALACTOSIDASE ON NATURAL AND SYNTHETIC POLYMERS TATAR ÖNAL, Seçil PhD in Biochemistry Supervisor: Prof. Dr. Azmi TELEFONCU October 2000, 190 pages a-Galactosidases (a-D-galactosid galactohydrolase, EC 3.2.1.22), are a common class of hydrolases which are widely distributed in nature. They have been isolated and purified with different techniques from various sources, such as plants, animals and microorganisms. These enzymes catalyses the hydrolysis of a-l,6-galactose bonds which found in galactooligosaccharides and polysaccharides. They are very useful enzymes, especially for elucidation of the biological functions of complex carbohydrates, for organic synthesis, for structural analysis, for medical purpose and in sugar industry. Because of these facts, the isolation, purification and characterization of enzyme from different sources has a great importance at last years. But in contrast this, there is only a few immobilization studies with this enzyme. In this study, a-galactosidase was isolated and partially purified from watermelon (Citrullus vulgaris) at first and then immobilized on natural and synthetic polymers by covalent coupling. Watermelons, as asource öf a-galactosidase are readily available and they are rich in a-galactosidase activity. The watermelon enzyme was immobilized on chitin and an ion exchange resin (Amberlite IRA-938). Various types of immobilization methods and polymers compared and then the characterization of immobilized enzyme was searched according to the chosen method and polymer. At first, the polymers were activated with l,l'-carbonyldiimidazole (CDI) and then the enzyme was immobilized directly or with a spacer arm (y-aminobutirycacid; GABA) to the polymer. According to results of this method physical and chemical c^acterization of the enzyme was made. Some parameters effecting to the enzyme activity (amount of protein, incubation time, pH, temperature, and substrate concentration) and stability of the immobilized enzyme (thermal, pH, storage and operational stability) were also searched. Beside of these, usebility of the enzyme was tested and the most useful reactor type for the hydrolysis of raffinose was determined. Key words : Amberlite IRA-938, Citrulhis vulgaris,, watermelon, a-galactosidase, immobilization, ion-exchange resin, chitin, isolation and purification of proteins, hydrolysis of raffinose
VII ABSTRACT IMMOBILIZATION OF WATERMELON (C1TRULLUS VULGARIS) a-GALACTOSIDASE ON NATURAL AND SYNTHETIC POLYMERS TATAR ÖNAL, Seçil PhD in Biochemistry Supervisor: Prof. Dr. Azmi TELEFONCU October 2000, 190 pages a-Galactosidases (a-D-galactosid galactohydrolase, EC 3.2.1.22), are a common class of hydrolases which are widely distributed in nature. They have been isolated and purified with different techniques from various sources, such as plants, animals and microorganisms. These enzymes catalyses the hydrolysis of a-l,6-galactose bonds which found in galactooligosaccharides and polysaccharides. They are very useful enzymes, especially for elucidation of the biological functions of complex carbohydrates, for organic synthesis, for structural analysis, for medical purpose and in sugar industry. Because of these facts, the isolation, purification and characterization of enzyme from different sources has a great importance at last years. But in contrast this, there is only a few immobilization studies with this enzyme. In this study, a-galactosidase was isolated and partially purified from watermelon (Citrullus vulgaris) at first and then immobilized on natural and synthetic polymers by covalent coupling. Watermelons, as asource öf a-galactosidase are readily available and they are rich in a-galactosidase activity. The watermelon enzyme was immobilized on chitin and an ion exchange resin (Amberlite IRA-938). Various types of immobilization methods and polymers compared and then the characterization of immobilized enzyme was searched according to the chosen method and polymer. At first, the polymers were activated with l,l'-carbonyldiimidazole (CDI) and then the enzyme was immobilized directly or with a spacer arm (y-aminobutirycacid; GABA) to the polymer. According to results of this method physical and chemical c^acterization of the enzyme was made. Some parameters effecting to the enzyme activity (amount of protein, incubation time, pH, temperature, and substrate concentration) and stability of the immobilized enzyme (thermal, pH, storage and operational stability) were also searched. Beside of these, usebility of the enzyme was tested and the most useful reactor type for the hydrolysis of raffinose was determined. Key words : Amberlite IRA-938, Citrulhis vulgaris,, watermelon, a-galactosidase, immobilization, ion-exchange resin, chitin, isolation and purification of proteins, hydrolysis of raffinose
Açıklama
Anahtar Kelimeler
Biyokimya, Biochemistry, Alfa-galaktosidaz, Alpha-galactosidase, Hidrolazlar, Hydrolases, Karpuz, Watermelon, Polimerler, Polymers, İmmobilizasyon, Immobilization
