透過聚醣陣列探討A型流感血凝素與糖的結合專一性

Abstract

流感病毒是一種致病性強的呼吸道病毒，主要分為A、B、C三種類型，其中A型流感病毒尤具高度變異性，易引發大規模流行。大多數人感染後僅出現輕微症状，並通常能自行康復，但對於新生兒、老年人和慢性病患者，流感病毒可能引發嚴重併發症，如肺炎等，甚至導致死亡。在感染宿主細胞的過程中，流感病毒表面的血凝素（Hemagglutinin, HA）扮演關鍵角色。HA蛋白與宿主細胞表面的糖結合，這一特性對病毒感染及疫苗研發至關重要。糖微陣列是一種高通量的生物分析技術，能夠在單一實驗中同時測量多種糖與蛋白質之間的相互作用。本研究利用實驗室自製的糖微陣列模擬宿主細胞表面的糖結構，研究不同糖結構對HA蛋白結合的影響，以探討流感病毒HA蛋白與不同糖類之間的結合專一性，從而為流感疫苗的設計提供更多資訊。我們調查了14種H1N1、3種H5N1、2種B型和1種H3N2的HA蛋白。結果顯示，部分H1N1 HA蛋白會與Poly-LacNAc且末端具有α2,6鍵結的Neu5Ac的糖結合。其中，H1N1 A/Beijing/262/1995的HA蛋白能識別α2,3和α2,6鍵結的Neu5Ac。而H5N1 A/Vietnam/1194/2004（禽流感病毒）基於其結構特性，對末端具有α2,3聯結的糖具有強結合力。然而，其他株的HA蛋白在不同實驗條件下，在糖微陣列上未顯示任何結合。除了改變實驗條件，我們也透過紅血球凝集實驗檢查HA蛋白的活性，以及透過七種凝集素再次確認糖微陣列的可行性。

Influenza viruses, particularly type A, are highly pathogenic and prone to causing global pandemics. While most cases are mild, the virus can cause severe symptoms like pneumonia and even death, especially in infants, the elderly, and those with chronic illnesses. The hemagglutinin (HA) glycoprotein on the virus surface binds to glycans on host cells, playing a crucial role in infection and vaccine development. In this study, I used a glycan microarray with 182 different glycans to mimic host cell glycan structures and investigate HA protein binding. This helped explore the binding specificity between various influenza virus HA proteins and glycans, providing insights for vaccine design. I examined HA proteins from different strains and subtypes (14 H1N1, 3 H5N1, 2 influenza B, and 1 H3N2). Results showed that some H1N1 HA proteins bind to glycans with LacNAc repeats and α2,6-linked Neu5Ac, and H1N1 A/Beijing/262/1995 could recognize Neu5Ac with both α2,3 and α2,6 linkages. The H5N1 A/Vietnam/1194/2004 HA protein exhibited strong binding affinity to α2,3-linked glycans due to its structural characteristics. However, other HA proteins did not show any binding under various experimental conditions. In addition to modify the experimental conditions, we also examined the activity of the HA protein through HA assays, and confirmed the feasibility of the glycan microarray using seven different lectins.