KÜTLE SPEKTROMETRESİ TEMELLİ PROTEOMİK YAKLAŞIMLAR KULLANILARAK KEFİRİN DETAYLI KARAKTERİZASYONU
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Tarih
2022-08
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info:eu-repo/semantics/openAccess
Özet
Kafkasya kökenli kadim içecek kefir çok eski tarihlerden beri bir sağlık kaynağı olarak görülmektedir. Son yıllarda yapılan çalışmalar da bunu desteklemektedir. Kefirin bu önemli özellikleri göstermesinin altında yatan mekanizmalar henüz aydınlatılabilmiş değildir. Bu çalışmada kefir üretimi ve depolanması süreçlerinde süt protein profillerinde meydana gelen değişimlerin kütle spektrometresi temelli-omik yaklaşımlar kullanılarak incelenmesi amaçlanmıştır. Ayrıca kefir mikroflorasından süte geçen kefir bazlı proteinlerin de tespiti çalışma kapsamında incelenmiştir. İlk olarak kefir üretimi gerçekleştirilmiştir. Kefirin üretim ve depolanması süreçlerinde çalışmada belirlenen periyotlardan kefir örnekleri alınmış ve süt proteinleri ekstrakte edilmiştir. Klasik bir proteomik yaklaşım kullanılarak Lys-C ve tripsin enzimleri kullanılarak peptitler üretilmiştir. Sonrasında peptitlerin fraksiyonlanması gerçekleştirilmiştir. Peptit içeren örnekler nLC-QExactive-Plus kütle spektrometresi ile analiz edilmiştir. Analiz sonucu elde edilen veriler MaxQuant yazılımı ile işlenmiş ve istatiksel analizler gerçekleştirilmiştir. Analizler sonucunda süt proteinlerinde meydana gelen değişimler gösterilmiştir. Ayrıca kefir mikroflorasından süte geçen proteinler ve bu proteinlerin işlevleri çalışmada gösterilmiştir. Elde edilen sonuçlara göre süt proteomunda 0-12 saat aralığında 3 adet protein, 0-24 saat aralığında 39 adet protein, 0-7 gün aralığında 41 adet protein ve son olarak 0-28 gün aralığında 42 adet proteinde anlamlı değişiklikler saptanmıştır. 24. Saatten itibaren 23 adet proteinde anlamlı bir değişiklik gözlemlenmiştir. Analizlerde 398 adet kefir temelli proteinin süte salındığı çalışmada tespit edilmiştir. Bu proteinlerin 141 adedi etiketsiz olarak çalışmada miktarlanabilmiştir. Bu sonuçların kefir üretiminde süt proteomunda meydana gelen değişimleri göstermesi açısından literatüre değerli katkılar sağlayacağı öngörülmektedir.
Kefir, an ancient beverage originating from the Caucasus, has been seen as a source of health since ancient times. Recent studies also support this. The mechanisms underlying kefir's display of these essential properties have not yet been elucidated. This study aims to examine the changes in milk protein profiles during kefir production and storage by using mass spectrometry-based-omics approaches. In addition, the detection of kefir-based proteins that pass into milk from kefir microflora was also investigated within the scope of the study. First, kefir production by using bovine milk was carried out. During the production and storage processes of kefir, kefir samples were taken from the periods determined in the study, and milk proteins were extracted. Peptides were produced using Lys-C and trypsin enzymes using a classical proteomics approach. The peptides were then fractionated. Samples containing peptides were analyzed by nLC-QExactive-Plus mass spectrometry. The data obtained as a result of the analysis were processed with the Maxquant software, and statistical analyses were performed. As a result of the study, the changes in milk proteins were shown. Inaddition, the proteins that pass into milk from kefir microflora and the functions of these proteins are shown in the study. According to the results, significant changes were found in the milk proteome in 3 proteins between 0-12 hours, 39 proteins between 0-24 hours, 41 proteins between 0-7 days, and finally, 42 proteins between 0-28 days. From the 24th hour, a significant change was observed in 23 proteins. In the analysis, it was determined in the study that 398 kefir-based proteins were released into milk. One hundred forty-one of these proteins could be quantified in the study by a label-free approach. These results are anticipated to provide valuable contributions to the literature showing the changes in the milk proteome in kefir production."
Kefir, an ancient beverage originating from the Caucasus, has been seen as a source of health since ancient times. Recent studies also support this. The mechanisms underlying kefir's display of these essential properties have not yet been elucidated. This study aims to examine the changes in milk protein profiles during kefir production and storage by using mass spectrometry-based-omics approaches. In addition, the detection of kefir-based proteins that pass into milk from kefir microflora was also investigated within the scope of the study. First, kefir production by using bovine milk was carried out. During the production and storage processes of kefir, kefir samples were taken from the periods determined in the study, and milk proteins were extracted. Peptides were produced using Lys-C and trypsin enzymes using a classical proteomics approach. The peptides were then fractionated. Samples containing peptides were analyzed by nLC-QExactive-Plus mass spectrometry. The data obtained as a result of the analysis were processed with the Maxquant software, and statistical analyses were performed. As a result of the study, the changes in milk proteins were shown. Inaddition, the proteins that pass into milk from kefir microflora and the functions of these proteins are shown in the study. According to the results, significant changes were found in the milk proteome in 3 proteins between 0-12 hours, 39 proteins between 0-24 hours, 41 proteins between 0-7 days, and finally, 42 proteins between 0-28 days. From the 24th hour, a significant change was observed in 23 proteins. In the analysis, it was determined in the study that 398 kefir-based proteins were released into milk. One hundred forty-one of these proteins could be quantified in the study by a label-free approach. These results are anticipated to provide valuable contributions to the literature showing the changes in the milk proteome in kefir production."
Açıklama
Anahtar Kelimeler
Kefir, proteomik, fosfoproteomik, kütle spektrometrisi., Kefir, proteomics, phosphoproteomics, mass spectrometry.