Dupont Didier, Lugand Damien, Rolet-Repecaud Odile, Degelaen Jacques
L'Institut National de la Recherche Agronomique (INRA), Unité de Recherches en Technologie et Analyses Laitières, Place du Champ de Foire, F-39800 Poligny, France.
J Agric Food Chem. 2007 Aug 22;55(17):6857-62. doi: 10.1021/jf070694w. Epub 2007 Jul 24.
Casein proteolysis can occur in milk during storage leading to its gelation. The two main proteolytic systems suspected to be involved are the plasmin and the proteases produced by psychrotrophic bacteria. The latter have been shown to cleave kappa-casein at the Phe105-Met106 bond. Although several techniques allow the determination of plasmin in milk, few rapid and easy-to-perform analytical techniques are available to check for bacterial proteolytic activity. This study presents the development of an inhibition ELISA allowing for the quantification of the kappa-casein intact at the Phe105-Met106 bond. It uses a monoclonal antibody specifically directed against this peptide bond that binds to the protein as long as the molecule's cleavage site is intact but not when it is cleaved. This simple technique allows for the rapid analysis of more than 20 samples within 3 h. Applied to commercial milks, this assay allowed for the detection of unstable milk.
酪蛋白水解在牛奶储存过程中可能发生，导致牛奶凝胶化。怀疑参与其中的两个主要蛋白水解系统是纤溶酶和嗜冷菌产生的蛋白酶。后者已被证明可在Phe105 - Met106键处切割κ-酪蛋白。虽然有几种技术可用于测定牛奶中的纤溶酶，但用于检测细菌蛋白水解活性的快速且易于操作的分析技术却很少。本研究介绍了一种抑制ELISA的开发，该方法可对Phe105 - Met106键处完整的κ-酪蛋白进行定量。它使用一种特异性针对该肽键的单克隆抗体，只要分子的切割位点完整，该抗体就会与蛋白质结合，而在切割时则不会结合。这种简单的技术可在3小时内快速分析20多个样品。应用于市售牛奶时，该检测方法可检测出不稳定的牛奶。
