Hessler R J, Blackwood R A, Brock T G, Francis J W, Harsh D M, Smolen J E
Department of Pediatrics, University of Michigan, Ann Arbor, USA.
J Leukoc Biol. 1998 Mar;63(3):331-6. doi: 10.1002/jlb.63.3.331.
The membrane fusion events observed during neutrophil degranulation are important aspects of the immunoregulatory system. In an attempt to understand the regulation of granule-plasma membrane fusion, we have begun characterizing human neutrophil cytosol for fusion activity, finding that 50% of the fusogenic activity could be attributed to members of the annexin family of proteins. The major non-annexin fusion activity (25% of the total cytosolic activity) was enriched by ion exchange chromatography after depletion of annexins by Ca2+-dependent phospholipid affinity chromatography. The fusion activity co-purified with a 10,14-kDa dimer identified as leukocyte L1 (which was non-fusogenic), along with an approximately 36-kDa protein. This protein was identified as glyceraldehyde-3-phosphate dehydrogenase (GAPDH) by amino-terminal sequencing, and the fusion activity was verified using commercially available GAPDH. GAPDH may play an important role in degranulation because it is as potent as annexin I on a mass basis and may constitute up to 25% of the total cytosolic fusion activity of the neutrophil.
在中性粒细胞脱颗粒过程中观察到的膜融合事件是免疫调节系统的重要方面。为了理解颗粒-质膜融合的调控机制，我们开始对人中性粒细胞胞质溶胶的融合活性进行表征，发现50%的融合活性可归因于膜联蛋白家族的蛋白质成员。在用钙离子依赖性磷脂亲和色谱法去除膜联蛋白后，通过离子交换色谱法富集了主要的非膜联蛋白融合活性（占总胞质活性的25%）。融合活性与一种被鉴定为白细胞L1（无融合活性）的10、14 kDa二聚体以及一种约36 kDa的蛋白质共同纯化。通过氨基末端测序将该蛋白质鉴定为甘油醛-3-磷酸脱氢酶（GAPDH），并使用市售的GAPDH验证了融合活性。GAPDH可能在脱颗粒过程中起重要作用，因为按质量计算它与膜联蛋白I一样有效，并且可能占中性粒细胞总胞质融合活性的25%。
