编码日本梨树(Pyrus pyrifolia )花粉类β-1,3-葡聚糖酶蛋白的cDNA克隆与核苷酸序列

构建了日本梨树(Pyrus pyrifolia)花粉cDNA文库,发现了一个编码花粉管分泌的类β-1,3-葡聚糖酶蛋白(BGN-1)的cDNA(bgn-1)并测定了其序列.bgn-1由1408 bp组成,包括47 bp的5'-非翻译区,由1194bp组成并编码了397个氨基酸的ORF和167 bp的3'-非翻译区.3'-非翻译区含有1个NUE(AATTAA)和3个似FUE的基序,分别位于1369～1374(AATTAA)、1320～1325、1327～1332(TTTTTA)和1363～1368(TTTGGA).bgn-1编码的蛋白(BGN-1)与DDBJ中的植物β-1,3-葡聚糖酶的序列相似性范围为37%～59%,与3D结构已知的大麦β-1,3-葡聚糖酶(GⅡ)的序列相似性为39.7%.对GⅡ与BGN-1进行的疏水簇分析(HCA同源分为87.1%,＞75%的一般标准)和使用3D-PSSM软件进行的分析暗示,BGN-1的3D结构与大麦GⅡ同源性最大(≥95%的肯定度).BGN-1的信号肽长度为22 aa,成熟的BGN-1(375 aa)的理论分子量为4 0723 D,理论pI值为9.59,诊断氨基酸残基模式(D,L,S,L)与禾谷类的与生长相关的亚家族D的诊断氨基酸残基模式(D,L,S,Q)极相似.在BGN-1的C-端延伸区的位点352～367之间发现了1个功能不清,重复出现的ProXXPro结构.

Cloning and Nucleotide Sequence of the cDNA Encoding a β -1,3-Glucanase-like Protein Secreted from Growing Pollen Tubes of Japanese Pear (Pyrus pyrifolia)

A pollen cDNA library of Japanese pear (Pyrus pyrifolia ) of the family Rosaceae was constructed and a cDNA (bgn-1)of 1 408 bp, which encodes a protein (BGN-1) secreted from growing pollen tubes, has been cloned and sequenced. bgn-1 cDNA is composed of a 5'-untranslated region of 47 bp, an open reading frame of 1 194 bp encoding 397 amino acid residues and a complete 3'-untranslated region of 167 bp. Alignment of the deduced amino acid sequence of bgn-1 with that of the barley (Hordeum vulgare)β -1 ,3-glucanase (G Ⅱ) showed 39.7% of amino acid identity. Several residues that were critical for the function of G Ⅱ were conserved in the deduced BGN-1 polypeptide. Moreover, a hydrophobic cluster analysis (HCA) showed an overall HCA homology score of 87.1% and analysis of BGN-1 with the 3D-PSSM program predicted a 3D structure of BGN-1 highly homologous to that of barley G Ⅱ with ≥ 95% certainty. These results suggest that the cloned bgn-1 cDNA encode a β -1,3-glucanase-like protein in Japanese pear.The predicted mature protein (375 aa) has a theoretical molecular mass of 40 723 D, a basic pI of 9.59, a diagnostic amino acid residue mode of D, L, S and L which is very similar to that of growth-related subfamily D (D, L, S and Q) in cereal. A ProXXPro repeat is also found between the position 352 and 367 in the C-extension region.