FAQ Peptide Synthesis

  1. The basic principle of peptide synthesis?
  2. Solid phase peptide synthesis is an important breakthrough in peptide synthesis chemistry. Its main feature is that it does not need to purify intermediate products, and the synthesis process can be carried out continuously, which lays a foundation for the automation of peptide synthesis. At present, the fully automatic synthesis of peptides is basically solid-phase synthesis. The basic process is as follows,

    Based on Fmoc chemical synthesis, the carboxyl group of the C-terminal amino acid of the target peptide is covalently linked with an insoluble polymer resin, and then the amino group of this amino acid is used as the starting point of peptide synthesis to form peptide bonds with the activated carboxyl groups of other amino acids. The peptide can be obtained by repeating this process. According to the different amino acid composition of peptides, the post-processing methods of peptides are different, and the purification methods are also different.

  3. How to control the quality of custom peptide services?
  4. BOC Sciences's peptide is manufactured in strict accordance with the ISO quality management system, with a unique numbered management for each peptide. The crude product, purified collection solution and final freeze-dried product are detected and analyzed by three-channel HPLC and MS methods to ensure the correctness and quality of the product.

  5. What methods are commonly used to produce peptides?
  6. The linear peptide’s peptide chain is extended by Fmoc solid-phase synthesis method, and amino acids are connected step by step from C-terminal to N-terminal. At the beginning, the first amino acid was linked to the insoluble support resin through an acid-sensitive linker. After removing the Fmoc protective group with hexahydropyridine, the second Fmoc protected amino acid is connected. The connection method includes pre-activation or "one-pot" and so on. After the target sequence was linked, the peptide chain was eluted from the resin with TFA to obtain the crude product.

  7. How to choose the purity of peptide for different experiments?
  8. The purity of peptide is a very important index, and the choice of purity depends on the purpose of the experiment. For the less sensitive screening test, it is recommended to use crude product or > 75%, and for the immune level, it is recommended to use > 85%. For the study of receptor-ligand interaction, biological assay, or cell level studies, the recommendation is >95%, and for the structural study, the recommendation is >98%.

  9. What is the net content of peptide (peptide content) and what is its significance?
  10. The weight of dry peptide not only contains peptide, but also contains some non-peptide components, such as water, absorbed solvent, coordination ions and salts. The net content of peptide refers to the weight percentage of peptide in it. The value of this percentage can range from 50% to 90%, depending on purity, sequence, and synthesis and purification. Do not confuse the net content of the peptide with the purity of the peptide. They are two completely different concepts. Purity is usually determined by HPLC. And purity is defined as the percentage of components with correct sequence in a peptide sample, while the net content of peptide refers to the percentage of peptide substances relative to non-peptide substances in the sample. The net content of peptide is usually determined by amino acid composition analysis or ultraviolet spectrophotometry. This information is very important to calculate the concentration of peptide in some experiments which are sensitive to the concentration of peptide.

  11. If the peptide purity is 95%, what are the remaining 5%?
  12. The purity of peptides is usually determined by HPLC with a standard acetonitrile gradient of 1% per minute. In the synthesis process, the cross-linking efficiency between amino acids can not always reach 100%, which resulting in a series of amino acid deletion impurities. Most of these amino acid-deficient impurities were removed during purification, but whose chromatographic performance is very similar to the target peptide. These amino acid deletions remained in the peptide sample for the remaining few percent.

  13. What impurities are present in non-HPLC purified peptides?
  14. Peptides and non-peptide impurities in crude and desalted grades: such as non-full-length peptides and some raw materials for peptide post-processing such as DTT, TFA, etc.

  15. How to calculate the molecular weight of peptide?
  16. The molecular weight of peptides can be calculated by some softwares such as peptide companion or by directly inputting the sequence on some websites. Due to the difference of decimal point in calculation, the decimal part of molecular weight may be different.

  17. What length of peptide is appropriate?
  18. Peptide synthesis requires consider factors such as the length, charge, and hydrophobicity of the peptide. The longer the length, the lower the purity and yield of the crude product, and the more difficult of purification and the greater the probability of failure to synthesize. Of course, the sequence of peptide functional region can’t be changed, but for the smooth synthesis of peptide, sometimes it is necessary to add some auxiliary amino acids in the upstream and downstream of the functional region to improve the solubility and hydrophilicity of peptide. If the peptide is too short, there may be problems in the synthesis. The main problem is that the synthesized peptide has a certain difficulty in the post-treatment process. The peptides below 5 peptides generally need hydrophobic amino acids, otherwise the post-treatment is more difficult. Peptides with less than 15 amino acid residues were obtained in satisfactory yield.

  19. How to determine the solubility of peptide from peptide sequence?
    1. If the peptide contains a high proportion of hydrophobic amino acids such as Leu, Val, IIe, Met, Phe and Trp, it is difficult to dissolve in aqueous solutions or impossible to dissolve at all.. These amino acids, whether purified or synthesized, may have problems.
    2. Generally the proportion of hydrophobic amino acids is < 50%, 5 consecutive amino acids can not be hydrophobic, and the proportion of charged amino acids (positive charge K, R, H, N-terminus, negative charge D, E, C - terminus) reaches 20%, in peptide N or C short can also improve solubility if polar amino acids can be increased.
  20. Why is it difficult to synthesize peptides containing Cys, Met, or Trp?
  21. It is difficult to synthesize peptides containing Cys, Met, or Trp and obtain high purity products. The main reason is that these groups are unstable and easy to oxidize. Special attention should be paid to the use and storage of these peptides to avoid repeated opening the lid.

  22. Why is the synthetic yield or purity of some peptides low?
  23. There is a relatively large difference between peptide synthesis and primer synthesis. There are few primer that can not be synthesized, but there are often peptides that can not be synthesized. As Val, Lle, Tyr, Phe, Trp, Leu, Gln, and Thr are amino acids more orthologous or repeated, the peptide chain can not be dissolved completely during the synthesis process and the synthesis efficiency decreases. In the following cases, the synthesis efficiency and purity of the product are relatively low, such as: repetitive pro, Ser-Ser, repetitive ASP, four continuous Gly, etc.

  24. How to purify the peptide?
  25. The peptide was purified by reversed-phase column (such as C8, C18, etc.) at 214 nm. The buffer system is usually a solvent containing TFA at pH 2.0. Buffer A contains 0.1% TFA in ddH2O, and Buffer B contains 1% TFA / ACN / ph2.0. Buffer A is used to dissolve the peptide before purification; if the solution is not good, Buffer B is used to dissolve it, and then Buffer A is used to dilute it. For strongly hydrophobic peptides, it is sometimes necessary to add small amounts of formic acid or acetic acid. HPLC analysis of crude peptide products, if the peptide is not long (below 15 aa), generally there will be a main peak, the main peak is usually full-length product; for the long peptide above 20 aa, if there is no main peak, HPLC needs to match mass to determine the molecular weight, and then determine which peak is the peptide to be synthesized.

  26. How long is the appropriate peptide for immunization?
  27. Generally it is about 10-15 amino acids. Of course the larger the number of amino acids, the better the immune effect, but the synthesis expense will also increase. Map peptides are then expected to be more than 15 aa in length. In addition, peptides under 10 aa were less effective in immunization.

  28. The solubility of the peptides we synthesized is not good, so there are problems with the peptides, right?
  29. It is difficult to predict accurately the solubility of a peptide and what a suitable solvent is. This notion is not true if peptides are difficult to solubilize and it is assumed that peptide synthesis is problematic.

  30. What are the appearance of peptides?
  31. The peptides we provide are powdered and generally white. The color of peptide powder varies with the composition, such as some yellow green with FITC modification.

  32. How to dissolve peptides?
  33. Dissolving peptides is very complicated, and it is generally difficult to determine the appropriate solvent at once. It is usually to take a little first for the test, and do not dissolve it completely before determining the suitable solvent.

    The following method helps you select the appropriate solvent:

    1. Determine the specific charge of a peptide, setting the acidic amino acids ASP (D), Glu (E) and C-terminal COOH as -1; the alkaline amino acids Lys (K), Arg (R), His (H) and N-terminal NH2 as +1, and the charges of other amino acids as 0. The number of charges was calculated.
    2. If the net charge number is > 0, the peptide is alkaline, dissolve with water: if not soluble or not very soluble, add acetic acid (10% more); if the peptide is not soluble yet, add a small amount of TFA (25 ul) to dissolve, and then add 500 ul of water to dilute.
    3. If net charge number is < 0, peptide is acidic, dissolve with water; if not dissolved or not very soluble, add ammonia (25 ul) to dissolve and then add 500 ul water to dilute.
    4. If the net charge number =0, the peptide is neutral, it generally needs to be dissolved with an organic solvent such as acetonitrile, methanol or isopropanol, DMSO, etc. It has also been suggested that urea is required to solubilize peptides of great hydrophobicity.
  34. How to store synthetic peptides?
  35. Peptides generally require protection from light for long-term storage and should be stored at –20℃ and at 4℃ for short-term storage. Peptide can be transported at room temperature for a short time. Peptides are stable at -20℃, especially freeze-dried and stored in a desiccator. Freeze dried peptides can be stored at room temperature before they are exposed to air. This will reduce the influence of humidity. When lyophilization is not possible, the best way is to store the sample in a small amount.

    For peptides containing Cys, Met or Trp, a deoxygenation buffer is essential for their solubilization because such peptides can be easily air oxidized. Nitrogen or argon, which slowly flows through the peptide before capping, also decreases oxidation. Peptides containing Gln or Asn are also easy to degrade, and all of these peptides have a limited life span compared with those without these problematic and simple peptides.

  36. How to deal with the two ends of peptide? Is it free or protected?
  37. Peptides are used to mimic proteins. In order to mimic the expression of proteins, we need to synthesize peptides with similar structure and charge to proteins. When a peptide is "cut out" from a protein, the number of charges at both ends will be different from that of the gene body protein. We need to change the synthesis strategy to make them consistent. In general, if it comes from the C-terminal of the protein, the N-terminal is shielded by acetylation; if it comes from the N-terminal of the protein, the C-terminal is shielded by amidation; if it comes from the middle part of the protein, both ends are shielded by acetylation and amidation.

  38. What is electrospray ionization mass spectrometry (electrospray ionization, EIS)?
  39. EIS can produce multivalent ionized proteins or peptides, and allow the analysis of proteins with a relative molecular weight of 1 × 105, with a resolution of 1500-2000 amu. The accuracy is about 0.01%. EIS is more suitable for on-line analysis of proteins with high molecular weight and needs gasification or organic solvents to sensitize the samples.

  40. What is matrix-assisted laser disso-ciation / ionization time of flight mass spectrometry (MALDI-TOF MS)?
  41. MALDI-TOF is currently a means of accurately determining molecular masses in protein identification and is particularly suitable for the determination of relative molecular masses of mixed protein peptide species with good sensitivity and resolution. It is an obligatory tool for current proteomic research. A hyphenated technique combining simultaneously with liquid chromatography can identify peptide species with high efficiency. Especially when the tandem application of various principles of mass spectrometry technology, not only can one obtain information on the relative molecular mass of a peptide but also determine its sequence structure, and this technique will be decisive in future proteomic studies.

  42. What is high performance liquid chromatography (HPLC)? What is reverse phase high performance liquid chromatography (RP-HPLC)?
  43. The appearance of HPLC provides a favorable method for the separation of peptides. Compared with other compounds, under suitable chromatographic conditions, the separation of proteins and peptides can be completed in a short time. What's more, HPLC can produce bio-active peptides on a large scale. Therefore, many scholars have done a lot of work to find the best conditions for the separation and preparation of peptides. How to maintain peptide activity, how to choose stationary phase materials, eluent types, how to analyze and determine are the current research contents.

    Reversed phase high performance liquid chromatography (RP-HPLC): the relationship between the results and retention value: to separate peptides by RP-HPLC, it is necessary to determine the retention of peptides with different structures on the column. In order to obtain a series of retention coefficients, Wilce et al. analyzed the retention properties and structures of 2,106 kinds of peptides by using multiple linear regression method, and obtained the relationship between different amino acid compositions and retention coefficients. The retention time of polar amino acid residues in 2-20 amino acid peptides can be reduced; in 10-60 amino acid peptides, more non-polar amino acids can also reduce the retention time on the column, while in 5-25 amino acid peptides, more non-polar amino acids can prolong the retention time on the column. At the same time, many literatures reported the effects of peptide chain length, amino acid composition, temperature and other conditions on the retention, and obtained the best conditions for the separation and extraction of each peptide by computer processing and analysis.

  44. Why is gradient elution used in RP-HPLC?
  45. At present, RP-HPLC is the most commonly used method to study the synthesis of peptide related substances. However, due to the different properties of related substances in synthetic peptides, it is difficult to fully detect all kinds of organic impurities in products by simple isocratic elution method.

  46. How to determine the monoisotopic peak and average isotopic peak in the mass spectrometry?
  47. In the mass spectrum with low resolution, the peak shown is generally the average isotope peak. However, the resolution of mass spectrometry (such as MALDI-TOF) is very high. The general small molecules (such as peptide segments) often contain 4-5 isotopic peaks, among which the smallest molecular weight is monoisotopic peak. When using mass spectrometry to detect macromolecules (such as intact protein), the number of isotopic peaks is very large, which is difficult to distinguish on the map. In this case, it makes more sense to use the average isotope peak.

  48. Is there a difference of 1 Da between the isotope peaks?
  49. Isotopic peptides generally differ by 1 Da in mass (m), but the peak detected by mass spectrometry is the mass charge ratio (m/z) of peptide rather than the mass itself, the mass charge ratio difference is less than 1 when the peptide has multiple charges. If the detected peptide has one charge, the difference between the isotopic peaks is 1; if the detected peptide has two charges, the difference between the isotopic peaks is 0.5; if the detected peptide has three charges, the difference between the isotopic peaks is 0.33; and so on. Therefore, from the distance between the isotopic peaks, we can infer the charged condition (z) of the peptide, and thus infer the single isotopic mass of the peptide [(m/z)*z].

  50. Is the monoisotopic peak necessarily the highest one?
  51. A monoisotopic peak must have the lowest mass charge ratio in a group of peaks, but not necessarily the highest. When the molecular weight of the peptide is small, the total number of atoms in the peptide is also small, the probability of the whole peptide containing isotope atoms is relatively small. At this time, the monoisotopic peak is often the highest peak. When the molecular weight of peptide increases, the probability of carrying isotope atoms increases. In this case, the isotope peak of +1 Da or even +2 Da may be the highest. The statistical results show that when the molecular weight of peptide is more than 2,500 Da, the peak is not a monoisotopic peak.

  52. Apart from HPLC and MS, what other peptide analysis methods are available?
  53. Water fraction, ion chromatography, elemental analysis, amino acid composition analysis, circular dichroism, NMR, IR, UV, endotoxin, microorganisms and other analyses can also be provided.

  54. What is the counter ion generally carried by peptides?
  55. Freeze-dried powder of peptide is generally provided with trifluoroacetate, if it is to do cell experiments or animal experiments, it is recommended to convert to acetate or hydrochloride.

  56. What isotope labeled peptides can be provided?
  57. Some stable isotope modified peptides, mainly including N15, C13 and deuterium modification can be provided. Since raw material of stable isotope amino acids are expensive, it is recommended to select some simple amino acids for labeling, such as Gly, Val, Phe, Leu, Ala, etc.

  58. What are the factors that affect the stability of peptides?
  59. Factors affecting the stability of synthetic peptides include deamidation, oxidation, hydrolysis, disulfide bond mismatch, racemization, β-elimination, aggregation, etc. The most common degradation products of synthetic peptides are deamidated products, oxidation products and hydrolysates. Among all kinds of amino acids, asparagine and glutamine are easy to deamidate (especially under high pH and high temperature); methionine, cysteine, histidine, tryptophan and tyrosine are most easily oxidized and sensitive to light; the peptide chain formed by aspartic acid is easy to break, especially ASP-Pro and ASP-Gly peptide bonds. The aggregation of peptides is mainly caused by hydrophobic interaction. Although at present it is difficult to accurately predict which peptides are prone to aggregation, at least for some medium and long peptides, it is necessary to study the possible existence of polymers.

  60. How to choose resin for Fmoc/tBu solid phase synthesis (SPPs)?
  61. For the synthesis of C-terminal carboxylic peptide, we can choose Wang resin; for the synthesis of C-terminal amide peptide, we can choose Rink Amide AM Resin or Rink Amide MBHA Resin; for the synthesis of full protection peptide, we can choose 2-Cl Trt Resin.

  62. What are the parameters of resin? What is the meaning of the parameters?
  63. Generally, the parameters of resin include loading, mesh number and specification, such as 1% DVB.
    Loading: the unit is mmol/g, that is, how many mmol of functional groups are present per gram of resin.
    Mesh: the particle size is usually 100-200 Mesh, the larger the value, the finer the particle.
    1% DVB: proportion of crosslinker divinylbenzene in styrene and divinylbenzene copolymers.

  64. What are the calculation methods of loading with Fmoc protected amino acid resin?
  65. There are generally two ways to calculate Fmoc protected amino acid resin loading, one is the weight gain method, where the increased weight is divided by the molecular weight and then divided by the total weight of the resin. The other is by UV analysis. At present, our company provides the loading value by UV analysis.

  66. What are the commonly used fluorescein modifications?
  67. DyeEx(nm)Em(nm)
    FITC488518
    5-FAM490520
    5-TAMRA544572
    TRITC547572
    Mca322390
    Amc351430
    AFC380500
    NBD466539
    Dnp350
    Texas Red589615
    Dabcyl426
    Glu(EDANS)335493
  68. What is FRET assay?
  69. Detection can be achieved using the principle of fluorescence resonance energy transfer (FRET), in which the fluorescent group EDNAS and the fluorescence quenching group DABCYL are labeled on both ends of a synthetic protease substrate peptide. EDNAS can emit fluorescence with a maximum emission wavelength of 490 nm upon excitation with a 355 nm light wave, while the quencher group DABCYL can absorb fluorescence with a maximum absorption wavelength of 490 nm as well. When the protease substrate is intact, the fluorescence emitted by the luminescent group is absorbed by the quenching group and the whole reaction system is not luminescent. When the protease substrate is cleaved, the luminescent group leaves the quenching group, the quenching effect decreases or even disappears, and the reaction system emits fluorescence.

  70. What are the common combinations of fluorescent groups and quenching groups?
  71. DABCYL/ EDANS,DABCYL/6-FAM,TAMRA/6-FAM,Abz/Tyr(3-NO2), Mca/Dnp.

  72. What are the ways of fluorescent modification in the C-terminal of the sequence?
  73. Usually, fluorescent dye compounds have carboxylic acid or NHS active ester, so amino functional group is needed when binding withpeptide. Therefore, we can modify the Lys side linked amino group in the sequence. Lys or ethylenediamine (ED: NH2CH2CH2NH2) were added to the C-terminal, such as Lys(Biotin),ED-Biotin,EDDnp, etc.

  74. What about the peptides modified by maps?
  75. Maps is a multi-antigen peptide, which is a branched peptide formed by linking the C-terminal of linear peptide to two or four Lys, thus increasing the size of the whole molecule. It usually has two, four or eight branches. When synthesizing maps peptides, the peptide products are qualitatively similar to non-target peptides due to heterogeneity upon condensation and thus are difficult to remove. HPLC purification is also difficult to provide mass spectrometric identification. Amino acid composition analysis is recommended.

  76. What are the effects of PEG modification?
  77. PEG modifier, also known as polyethylene glycol modifier, modified PEG, PEG modifier, is a kind of polyethylene glycol with functional groups. At present, it is mainly used for protein drug modification to increase the half-life in vivo, reduce immunogenicity, and increase the water solubility of drugs. In recent years, modified PEG has been widely used in drug discovery and development, and plays an important role in drug sustained release.

  78. What are the ways of PEG modification?
  79. The common modification groups of PEG modifier can be summarized as follows:
    Amino(-amine)-NH2, aminomethyl-CH2-NH2, maleimide-Mal, carboxyl-COOH, sulfhydryl(-thiol)-SH, succinimide carbonate-SC, succinimide acetate-SCM, succinimide propionate-SPA, succinimide-NHS, propionyl-CH2CH2COOH, aldehyde-CHO, acrylate-AC, azide, biotin, fluorescein, glutaric acid-GA, hydrazide, alkyne, etc.
    The common classifications of PEG modifiers are as follows

    1. Linear PEG modifier: mPEG
    2. Bifunctional modifiers: HCOO-PEG-COOH,NH2-PEG-NH2,OH-PEG-COOH,OH-PEG-NH2,HCL·NH2-PEG-COOH,MAL-PEG-NHS
    3. According to the molecular weight, there are 2,000, 3,000, 5000, 10,000, 20,000, etc

    Therefore, it is necessary to determine the type of PEG, functional group requirements and molecular weight range in PEG modification.

  80. What is small molecule PEG?
  81. Small molecule PEG is a kind of compound with small molecular weight (such as mini-PEG), definite molecular weight, amino group at one end and carboxylic acid at the other. The following compounds are commonly used.

    AEA5-amino-3-oxapentanoic acid
    AEEA8-Amino-3,6-Dioxaoctanoic Acid   9 atoms (mini-PEG)
    TTDS1,13-diamino-4,7,10-trioxatridecan-succinamic acid
    Ebes8 - amino - 3,6 - dioxa - octyl)succinamic acid (PEG2-Suc-OH)
    AEEEA11-Amino-3,6,9-Trioxaundecanoic Acid  11 atoms(mini-PEG3)
    dPEG(4)15-amino-4,7,10,13-tetraoxa-pentadecanoic acid
    dPEG(8)alpha-amino-omega-carboxy octa(ethylene glycol)
    dPEG(12)alpha-amino-omega-carboxy dodeca(ethylene glycol)
  82. Can pseudoproline dipeptides help the synthesis of difficult peptides?
  83. Yes, pseudoproline dipeptides can reduce the alpha angle and beta folding space structure of peptide in peptide condensation, keep the peptide linear, facilitate the synthesis of peptide and improve the purity of crude peptide. The structure of pseudoproline is removed when the peptide is cleaved with TFA, and the normal peptide is obtained. Pseudoproline dipeptides mainly consists of two major series, Fmoc-AA-Ser(Psi(Me,Me)pro)-OH and Fmoc-AA-Thr(Psi(Me,Me)pro)-OH, AA represents amino acids. If there is a side chain, the side chain should be protected.

    For example, Fmoc-Ser(tBu)-Ser(Psi(Me,Me)pro)-OH and Fmoc-Asp(OtBu)-Thr(Psi(Me,Me)pro)-OH.

  84. What are the Antibodies to Epitope Tags in common antibodies?
  85. TagSequence
    HISHHHHHH
    c-MYCEQKLISEEDL
    HAYPYDVPDYA
    VSV-GYTDIEMNRLGK
    HSVQPELAPEDPED
    V5GKPIPNPLLGLDST
    FLAGDYKDDDDK
  86. What is TAT peptide?
  87. The sequence of TAT peptide is Tyr-Gly-Arg-Lys-Lys-Arg-Arg-Gln-Arg-Arg-Arg (RKKRRORRR), which can carry peptide through membrane.

  88. What kind of peptides can be used to calculate the peptide content by UV analysis?
  89. If there is Trp or Tyr in the peptide sequence, the content of peptide can be analyzed by ultraviolet method, according to the molar extinction coefficient:
    Tryptophan 5,560 AU/mmole/ml
    Tyrosine 1,200 AU/mmole/ml
    (at 280 nm at neutral pH using a 1 cm cell)
    The molar extinction coefficient of a peptide can be added according to the coefficients of each Trp or Tyr, and then calculated by the formula according to the measured absorption value at 280 nm.
    mg peptide per ml = (A280 x DF x MW) / e
    A280, the actual absorption value (1 cm cell) at 280 nm,
    DF, dilution factor,
    MW, molecular weight of peptide
    e, molar extinction coefficient of peptide

  90. What modifications are available for click peptide?
  91. The representative reaction of click chemistry is copper catalyzed azide alkyne cycloaddition. The concept of click chemistry has made a great contribution to the field of chemical synthesis. In many fields such as drug development and biomedical materials, it has become one of the most useful and attractive synthesis concepts. It is mainly used in peptides with azido or alkynyl functional groups. The common compounds are as follows.
    Fmoc–4-azidophenylalanine   
    Fmoc-Phe(N3)–OH;
    Fmoc–Azidohomoalanine
    Fmoc-D–propargylglycine  
    Fmoc-D-Pra-OH
    Fmoc-L-propargylglycine   
    Fmoc-L-Pra-OH
    Fmoc-Lys(N3)-OH
    Fmoc-azidolysine
    Fmoc-lys(azide);
    Azidoacetic acid
    6-Azidohexanoic acid
    Propiolic acid

  92. What are the side chain methyl modified amino acids?
  93. They are Lys(Me), Lys(Me2), Lys(Me3), Orn(Me), Orn(Me2), Orn(Me3), Arg(Me), Arg(Me2,Symetrical), Arg(Me2,ASymetrical).

  94. What are the applications of C-terminal thioester peptides?
  95. The C-terminal thioester modification of peptide can be applied to the native chemical ligation reaction, which can undergo chemoselective transthioesterification reaction with another N-terminal cysteine peptide under neutral conditions, and rearrange into amide bond to realize the connection between the two peptides. It can be applied to the ligation of long peptides by fragments.

  96. What factors should be considered for peptide binding to other compounds or carriers?
  97. Some peptides often need to connect with some drugs or compounds with specific functions, or to some carriers. These processes need chemical bonds to combine. Generally, when peptides have amino or carboxylic acid functional groups, the corresponding carboxylic acid or amino group can be considered for condensation. Another option is to connect Maleimide with thiol in the environment of pH 8. The peptides can be modified with Cys or Maleimide functional groups. Maleimide modification can usually be achieved by 3-Maleimidopropionic Acid (CAS 7423-55-4).

  98. How to convert units?
  99. 1mg = 1,000 mcg (or μg)
    1ml=1,000 uL
    1 mol = 1,000 mmol
    1 mmol = 1,000 μmol
    1 μmol = 1,000 nmol
    1 nmol = 1,000 pmol

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