ABSTRACT
3-Hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR) plays an important role in catalyzing the first committed step of isoprenoids biosynthesis in mevalonic acid (MVA) pathway. Here, we cloned a full-length transcript of Paris fargesii Franch. The full-length cDNA of P. fargesii HMGR (Pf-HMGR, GenBank accession no. JX508638) was 1,973 bp and contained a 1,728 bp ORF encoding 576 amino acids. Sequence analysis revealed that the deduced Pf-HMGR had high similarity with HMGRs from other plants, including Ricinus communis (77%), Litchi chinensis (76%), Michelia chapensis (75%) and Panax quinquefolius (72%). It had a calculated molecular mass of about 62.13 kDa and an isoelectric point (pI) of 8.47. It contained two transmembrane domains, two putative HMGR binding sites and two NADP(H)-binding sites. The predicted 3-D structure revealed that Pf-HMGR had a similar spatial structure with other plant HMGRs. Three catalytic regions, including L-domain, N-domain and S-domain were detected by structural modeling of HMGR. Tissue expression analysis revealed that Pf-HMGR was strongly expressed in roots and stems than in leaves. Taken together, our data laid a foundation for further investigation of HMGR's functions and regulatory mechanisms in plants.
Subject(s)
Amino Acid Sequence , Base Sequence , Cloning, Molecular , Computational Biology , Hydroxymethylglutaryl CoA Reductases/chemistry , Hydroxymethylglutaryl CoA Reductases/genetics , Hydroxymethylglutaryl CoA Reductases/metabolism , Liliaceae/enzymology , Models, Molecular , Molecular Sequence Data , Protein Conformation , /genetics , Real-Time Polymerase Chain Reaction , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
Background: To observe the impact of application of bio‑amniotic membrane immersed in 5‑fluorouracil solution in trabeculectomy on the retina in a rabbit model. Materials and Methods: Healthy white New Zealand rabbits were randomly assigned into three groups with 20 in each group. Bio‑amniotic membranes of 4 × 5 mm immersed in either physiological saline/water for 10 min, or 25 mg/mL 5‑fluorouracil solution for 5 and 10 min, respectively, were applied on rabbit eyes during trabeculectomy. At 7, 14, 21, and 28 days of postoperation, five rabbits from each group were examined with electroretinogram (ERG). After being examined for eye pressure and bleb morphology, rabbits were sacrificed by air embolism and their retinas were collected and examined by transmission electron microscopy (TEM). In addition, 5‑fluorouracil amount in bio‑amniotic membranes was measured using high‑performance liquid chromatography. Results: Each bio‑amniotic membrane could absorb 59.004 μg and 75.828 μg 5‑fluorouracil after being immersed in 5‑fluorouracil solution for 5 and 10 min, respectively. Application of these bio‑amniotic membranes in trabeculectomy could promote the formation of well‑functioning bleb and maintain intraocular pressure, although it had no effect on retina structures as examined with ERG and TEM. Conclusion: Application of 5‑FU soaked bio‑amniotic membrane in rabbit eye trabeculectomy is effective and safe.