کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
1178564 | 962701 | 2006 | 5 صفحه PDF | دانلود رایگان |
![عکس صفحه اول مقاله: C-terminal residues of plant glutamate decarboxylase are required for oligomerization of a high-molecular weight complex and for activation by calcium/calmodulin C-terminal residues of plant glutamate decarboxylase are required for oligomerization of a high-molecular weight complex and for activation by calcium/calmodulin](/preview/png/1178564.png)
Bacterial glutamate decarboxylase (GAD) is a homohexameric enzyme of about 330 kDa. Plant GAD differs from the bacterial enzyme in having a C-terminal extension of 33 amino acids within which resides a calmodulin (CaM)-binding domain. In order to assess the role of the C-terminal extension in the formation of GAD complexes and in activation by Ca2+/CaM, we examined complexes formed with the purified full-length recombinant petunia GAD expressed in E. coli, and with a 9 amino acid C-terminal deletion mutant (GADΔC9). Size exclusion chromatography revealed that the full-length GAD formed complexes of about 580 kDa and 300 kDa in the absence of Ca2+/CaM, whereas in the presence of Ca2+/CaM all complexes shifted to ∼680 kDa. With deletion of 9 amino acids from the C-terminus (KKKKTNRVC500), the ability to bind CaM in the presence of Ca2+, and to purify it by CaM-affinity chromatography was retained, but the formation of GAD complexes larger than 340 kDa and enzyme activation by Ca2+/CaM were completely abolished. Hence, responsiveness to Ca2+/CaM is associated with the formation of protein complexes of 680 kDa, and requires some or all of the nine C-terminal amino acid residues. We suggest that evolution of plant GAD from a bacterial ancestral enzyme involved the formation of higher molecular weight complexes required for activation by Ca2+/CaM.
Journal: Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics - Volume 1764, Issue 5, May 2006, Pages 872–876