 Chemical Structure.png)
Physicochemical Properties
| Molecular Formula | C2H5NO |
| Molecular Weight | 59.067200422287 |
| CAS # | 9015-82-1 |
| Appearance | Typically exists as Off-white to light yellow solid at room temperature |
| Synonyms | Kininase II; CD143; Carboxycathepsin; Carboxypeptidase Zace2; Dipeptidyl carboxypeptidase; Dipeptidyl carboxypeptidase A; Dipeptidyl carboxypeptidase I; Dipeptidyl serine carboxypeptidase; E.C. 3.4.15.1; Native Porcine Angiotensin Converting Enzyme |
| Storage |
Powder-20°C 3 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition | Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs) |
Biological Activity
| ln Vitro |
ACE was discovered in the mid-1950s by the observation that dialysis of plasma and kidney extract with water and saline before incubation had produced two separate pressor substances, Ang I and Ang II respectively. It was discovered for a second time in 1966 during the characterization of bradykinin (BK) degrading enzyme from kidney and this enzyme was named kininase II; it later was found to be the same enzyme as ACE. ACE2 was discovered in 2000 when two independent research groups cloned homologous ACE that could convert Ang I to Ang(1–9) and yet also is captopril-insensitive [1]. Two isozymes of ACE are present in mammals: somatic ACE and testis ACE. Somatic ACE possesses two catalytic domains (N- and C-domains) and a C-terminal transmembrane segment (stalk) (Figure 29D.1 ). Somatic and testis ACEs in humans contain 1,306 and 665 aa residues, respectively. Testis ACE only possesses one catalytic domain. Both catalytic domains are zinc-metallopeptidase with the active motif HEMGH where the two histidine residues coordinate the zinc ion. The stalk anchors the enzyme on the membrane and is susceptible to be cleaved by shedding enzymes, resulting in plasma ACE activity (Figure 29D.1). ACE2 is a chimaera protein with a single catalytic domain of ACE, and a C-terminal highly resembling collectrin, which may act as a chaperone protein to deliver other proteins to the brush border membrane.[1] |
| ln Vivo |
Clinical Implications[1] Inclusion (II) or deletion (DD) of 287 bp Alu repeats in the 16th intron affects the human plasma ACE levels and the DD genotype was more frequently found in patients with myocardial infarction but no convincing evidence was available on the association of the DD genotype with hypertension. ACE2 was identified as the receptor for SARS (severe acute respiratory syndrome) coronavirus. SARS virus binding downregulates the cellular expression of ACE2, and the binding induces clathrin-dependent internalization of virus/receptor (SARS/ACE2) complex. Not only has ACE2 facilitated the invasion of SARS virus for rapid replication, but also ACE2 is depleted from the cell membrane and therefore the damaging effects of Ang II are enhanced, resulting in acute deterioration of lung tissues. Use for Diagnosis and Treatment[1] ACE has been the target of hypertension control since the 1970s. ACE inhibitors are prescribed as the sole or combinational treatment of high blood pressure, for the dual effects of lowering Ang II and slowing down BK degradation. In human hypertensive patients, ACE2 levels are lower in both kidney and heart compared to normotensive volunteers. |
| Enzyme Assay |
Gene and mRNA[1] ACE and ACE2 genes are located at chromosome 17q23 and Xp22 in humans, respectively. Testis ACE is transcribed from the same gene with an alternative transcription starting site on the 13th intron of the ACE gene, resulting in only C-domain and stalk segment with a unique additional 67 aa N-terminal sequence in humans. The two catalytic domains are the result of gene/domain duplication and the duplication occurred multiple times in evolution as the cnidarians, crustaceans, insects, and vertebrates possess ACE-like enzymes with one or two catalytic domains. No expression studies so far have been performed for non-mammalian ACE and ACE2. Distribution of mRNA[1] Somatic ACE is expressed in various tissues including blood vessels, kidney, intestine, adrenal gland, liver, and uterus, and is especially abundant in highly vascular organs such as retina and lung. Testis ACE is expressed by postmeiotic male germ cells and high-level expression is found in round and elongated spermatids. ACE2 is expressed in lung, liver, intestine, brain, testis, heart, and kidney. Regulation of Synthesis and Release[1] Expression of ACE is affected by steroids and thyroid hormone, but the details of the regulation are not clear. ACE is under promoter regulation by hypoxia-inducing factor 1α (HIF-1α), which upregulates the ACE expression under hypoxic conditions, resulting in an increase in Ang II concentration. Under hypoxia, ACE2 will be downregulated but it was shown that it is indirectly controlled by Ang II, but not HIF-1α. Testis ACE expression control is highly specific and regulated by a tissue-specific promoter located immediately −59 bp of the transcription start site, which is frequently used in testis-specific overexpression studies. Hypoxia induced by high temperature decreased gill ACE activity but had no effect on kidney in carp. Promoters of ACE2 from mammals, amphibians, and teleosts drive specific expression in the heart. Cis-element search results discovered WGATAR motifs in all putative ACE2 promoters from different vertebrates, suggesting a possible role of GATA family transcriptional factors in ACE2 expression regulation. |
| Cell Assay |
Target Cells/Tissues and Functions[1] The well-known function of ACE is the conversion of Ang I to Ang II and degradation of BK, which all play an important role in controlling blood pressure. ACE also acts on other natural substrates including encephalin, neurotensin, and substance P. Besides being involved in blood pressure control, ACE possesses widespread functions including renal development, male fertility, hematopoiesis, erythropoiesis, myelopoiesis, and immune responses. ACE2 can convert Ang II to Ang(1–7), thereby reducing the concentration of Ang II and increasing that of Ang(1–7). ACE2 can also convert Ang I to Ang(1–9), which is subsequently converted into Ang(1–7) by ACE. The high expression of ACE2 favors the balance of Ang(1–7) over Ang II, which accounts for the cardioprotective role of ACE2 via the Ang(1–7)/Mas signaling pathway. Tissue and Plasma Concentrations[1] Lung possesses the highest amount of ACE and contributes to 0.1% of total protein. Serum ACE levels in humans ranged from 299.3±49 μg/l (DD) to 494.1±88.3 μg/l (II) with heterozygous individuals 392.6±66.8 μg/l. (ID: see the section “Pathophysiological Implications” for the genotype definition.) Several enzymatic assays have been developed for the measurement of ACE activity in plasma and tissues and usually involve artificial substrates such as hippuryl-His-Leu or N-[3-(2-furyl)acryloyl]L-phenylalanyl-glycyl-glycine (FAPGG), in combination with captopril inhibition. These methods were developed in mammals but were also extended to other vertebrates including birds, amphibians, and fishes. However, these enzymatic methods may be erroneous because the enzyme specificity on the artificial substrates could be different. Lamprey ACE activities in different tissues were measured but captopril failed to decrease the ACE activities, indicating a possible nonspecific enzyme measurement. In amphibian, high captopril-sensitive ACE activities were found in gonad, intestine, kidney, and lung, moderate activities were presented in liver, heart, and skin, and low or negligible activities were observed in plasma, muscle, and erythrocytes. |
| Animal Protocol |
Phenotype in Gene-Modified Animals[1] ACE-knockout mice display normal blood pressure under normal conditions, but are sensitive to changes in blood pressure such as exercise. ACE-knockout also affects renal function, renal development, serum and urine electrolyte composition, haematocrit, and male reproductive capacity. Deficiency in testis ACE affects male fertility but its exact role is still not clear. Although mice with testis ACE deficiency mate normally and their sperm quantity and motility are no different from those of wild-type mice, the survival of sperm in the oviduct and fertilization rate are highly reduced. Overexpression of ACE2 in hypertensive models, but not in normotensive animals, reduced blood pressure. ACE2-knockout mice displayed progressive cardiac dysfunction resembling that of long-term hypoxia after coronary artery disease or bypass surgery in human, which could be reversed by concurrent ACE-knockout. It was suggested that the cardioprotective function of ACE2 is to counterbalance the effects of ACE. |
| References | [1]. Angiotensin Converting Enzymes. Handbook of Hormones. 2016:263–e29D-4. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7150253/ |
| Additional Infomation |
Angiotensin converting enzyme (ACE) is well known for its dual actions in converting inactive Ang I to active Ang II and degrade active bradykinin (BK), which play an important role in the control of blood pressure. Since the bottle neck step is the production of pressor Ang II, this was targeted pharmacologically in 1970s and successful ACE inhibitors such as captopril were produced to treat hypertension. Researches on domain specific ACE inhibitors are continuing to produce effective hypertension controlling drugs with fewer side effects. ACE2 was discovered in 2000; it converts Ang II into Ang(1–7), thereby reducing the concentration of Ang II as well as increasing that of Ang(1–7), an important enzyme for Ang(1–7)/Mas receptor signaling. ACE2 also acts as the receptor in the lung for the coronavirus causing the infamous severe acute respiratory syndrome (SARS) in 2003.[1] The first ACE inhibitor was a peptide antagonist called SQ 20,881 (GWPRPEIPP) discovered from snake venom but it was not orally active. The snake venom peptides were further studied to produce the first orally active form, captopril, that lowers the blood pressure of essential hypertensive patients. The most common side effects of captopril are cough, skin rash, and loss of taste, and therefore derivatives such as enalapril, lisinopril, and ramipril were developed with fewer side effects. After the discovery of N- and C-domains of ACE, specific domain inhibitors were developed to increase specificity. Ang I is mainly hydrolyzed by the C-domain in vivo but BK is hydrolyzed by both domains. By developing a C-domain selective inhibitor (RXPA380) some degradation of BK by the N-domain would be permitted and this degradation could be enough to prevent accumulation of excess BK causing angioedema.[1] |
Solubility Data
| Solubility (In Vitro) | May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples |
| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples. Injection Formulations (e.g. IP/IV/IM/SC) Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] *Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline) Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline) Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline) Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil) Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Oral Formulations Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). Oral Formulation 3: Dissolved in PEG400 Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose Oral Formulation 6: Mixing with food powders Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 16.9291 mL | 84.6453 mL | 169.2907 mL | |
| 5 mM | 3.3858 mL | 16.9291 mL | 33.8581 mL | |
| 10 mM | 1.6929 mL | 8.4645 mL | 16.9291 mL |