Instituto de Productos Lácteos de Asturias (IPLA-CSIC), Apdo, Asturias, Spain.
BMC Microbiol. 2011 Jun 17;11:138. doi: 10.1186/1471-2180-11-138.
Staphylococcus aureus is a food-borne pathogen and the most common cause of infections in hospitalized patients. The increase in the resistance of this pathogen to antibacterials has made necessary the development of new anti-staphylococcal agents. In this context, bacteriophage lytic enzymes such as endolysins and structural peptidoglycan (PG) hydrolases have received considerable attention as possible antimicrobials against gram-positive bacteria.
S. aureus bacteriophage vB_SauS-phiIPLA88 (phiIPLA88) contains a virion-associated muralytic enzyme (HydH5) encoded by orf58, which is located in the morphogenetic module. Comparative bioinformatic analysis revealed that HydH5 significantly resembled other peptidoglycan hydrolases encoded by staphylococcal phages. The protein consists of 634 amino acid residues. Two putative lytic domains were identified: an N-terminal CHAP (cysteine, histidine-dependent amidohydrolase/peptidase) domain (135 amino acid residues), and a C-terminal LYZ2 (lysozyme subfamily 2) domain (147 amino acid residues). These domains were also found when a predicted three-dimensional structure of HydH5 was made which provided the basis for deletion analysis. The complete HydH5 protein and truncated proteins containing only each catalytic domain were overproduced in E. coli and purified from inclusion bodies by subsequent refolding. Truncated and full-length HydH5 proteins were all able to bind and lyse S. aureus Sa9 cells as shown by binding assays, zymogram analyses and CFU reduction analysis. HydH5 demonstrated high antibiotic activity against early exponential cells, at 45°C and in the absence of divalent cations (Ca2+, Mg2+, Mn2+). Thermostability assays showed that HydH5 retained 72% of its activity after 5 min at 100°C.
The virion-associated PG hydrolase HydH5 has lytic activity against S. aureus, which makes it attractive as antimicrobial for food biopreservation and anti-staphylococcal therapy.
金黄色葡萄球菌是一种食源性病原体，也是住院患者感染的最常见原因。该病原体对抗菌药物的耐药性增加，使得开发新的抗葡萄球菌药物成为必要。在这种情况下，噬菌体裂解酶，如内切酶和结构肽聚糖（PG）水解酶，作为抗革兰氏阳性菌的潜在抗菌剂受到了相当多的关注。
金黄色葡萄球菌噬菌体 vB_SauS-phiIPLA88（phiIPLA88）含有一种由 orf58 编码的病毒相关溶壁酶（HydH5），该酶位于形态发生模块中。比较生物信息学分析表明，HydH5 与金黄色葡萄球菌噬菌体编码的其他肽聚糖水解酶有很大的相似性。该蛋白由 634 个氨基酸残基组成。鉴定出两个推定的裂解结构域：一个 N 端 CHAP（半胱氨酸、组氨酸依赖性氨水解酶/肽酶）结构域（135 个氨基酸残基）和一个 C 端 LYZ2（溶菌酶亚家族 2）结构域（147 个氨基酸残基）。当制作 HydH5 的预测三维结构时也发现了这些结构域，这为缺失分析提供了基础。完整的 HydH5 蛋白和仅包含每个催化结构域的截短蛋白在大肠杆菌中大量表达，并通过随后的复性从包涵体中纯化。截短和全长 HydH5 蛋白都能够结合并裂解 S. aureus Sa9 细胞，如结合测定、同工酶分析和 CFU 减少分析所示。HydH5 对早期指数细胞表现出高抗生素活性，在 45°C 且不存在二价阳离子（Ca2+、Mg2+、Mn2+）的情况下。热稳定性测定表明，HydH5 在 100°C 下 5 分钟后保留了 72%的活性。
病毒相关 PG 水解酶 HydH5 对金黄色葡萄球菌具有裂解活性，使其成为食品生物保鲜和抗葡萄球菌治疗的有吸引力的抗菌剂。
