Intracellular Antimicrobial Peptides Targeting the Protein Synthesis Machinery

  • Michael Graf
  • Daniel N. WilsonEmail author
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 1117)


While antimicrobial peptides (AMPs) are well-known for their disruptive effects on bacterial membranes, the mechanism of many intracellular AMPs is still being elucidated. In the recent years, it has been demonstrated that the subclass of proline-rich AMPs (PrAMPs) can pass through the bacterial membrane and kill bacteria by inhibiting protein synthesis. PrAMPs are a product of the innate immune system and are secreted in response to bacterial infection. So far PrAMPs have been identified in many arthropods, such as beetles, wasps, and flies, as well as some mammals, such as sheep, cows, and goats. PrAMPs show high potency against Gram-negative bacteria, while exhibiting low toxicity in eukaryotes, suggesting that they may represent a promising avenue for the development of future antimicrobial agents to combat the increase of multidrug-resistant bacterial pathogens. Structural and biochemical data have revealed the PrAMP binding sites on the ribosome as well as insight into their mechanisms of action. While the binding site of all so far investigated PrAMPs is situated within nascent polypeptide exit tunnel, the mechanism of action is distinct between class I and II PrAMPs. Specifically, class I PrAMPs, such as Bac7, Onc112, pyrrhocoricin, and metalnikowin, block the delivery of aa-tRNA by EF-Tu to the ribosomal A-site, whereas the class II PrAMPs, such as apidaecin 1b and Api137, act during translation termination and inhibit protein synthesis by trapping of release factors on the 70S ribosome following hydrolysis of the nascent polypeptide chain.


Proline-rich antimicrobial peptides Protein synthesis Translation Ribosome 


  1. Agerberth B, Lee JY, Bergman T, Carlquist M, Boman HG, Mutt V, Jornvall H (1991) Amino acid sequence of PR-39. Isolation from pig intestine of a new member of the family of proline-arginine-rich antibacterial peptides. Eur J Biochem 202:849–854CrossRefGoogle Scholar
  2. Asai T, Zaporojets D, Squires C, Squires CL (1999) An Escherichia coli strain with all chromosomal rRNA operons inactivated: complete exchange of rRNA genes between bacteria. Proc Natl Acad Sci U S A 96:1971–1976CrossRefGoogle Scholar
  3. Benincasa M, Scocchi M, Podda E, Skerlavaj B, Dolzani L, Gennaro R (2004) Antimicrobial activity of Bac7 fragments against drug-resistant clinical isolates. Peptides 25:2055–2061. CrossRefPubMedGoogle Scholar
  4. Berthold N et al (2013) Novel apidaecin 1b analogs with superior serum stabilities for treatment of infections by gram-negative pathogens. Antimicrob Agents Chemother 57:402–409. CrossRefPubMedPubMedCentralGoogle Scholar
  5. Brogden KA (2005) Antimicrobial peptides: pore formers or metabolic inhibitors in bacteria? Nat Rev Microbiol 3:238–250. CrossRefPubMedGoogle Scholar
  6. Bulet P et al (1993) A novel inducible antibacterial peptide of Drosophila carries an O-glycosylated substitution. J Biol Chem 268:14893–14897PubMedGoogle Scholar
  7. Casteels P, Tempst P (1994) Apidaecin-type peptide antibiotics function through a non-poreforming mechanism involving stereospecificity. Biochem Biophys Res Commun 199:339–345. CrossRefPubMedGoogle Scholar
  8. Casteels P, Ampe C, Jacobs F, Vaeck M, Tempst P (1989) Apidaecins: antibacterial peptides from honeybees. EMBO J 8:2387–2391CrossRefGoogle Scholar
  9. Casteels P et al (1990) Isolation and characterization of abaecin, a major antibacterial response peptide in the honeybee (Apis mellifera). Eur J Biochem 187:381–386CrossRefGoogle Scholar
  10. Casteels-Josson K, Capaci T, Casteels P, Tempst P (1993) Apidaecin multipeptide precursor structure: a putative mechanism for amplification of the insect antibacterial response. EMBO J 12:1569–1578CrossRefGoogle Scholar
  11. Castle M, Nazarian A, Yi SS, Tempst P (1999) Lethal effects of apidaecin on Escherichia coli involve sequential molecular interactions with diverse targets. J Biol Chem 274:32555–32564CrossRefGoogle Scholar
  12. Chernysh S, Cociancich S, Briand JP, Hetru C, Bulet P (1996) The inducible antibacterial peptides of the Hemipteran insect Palomena prasina: identification of a unique family of prolinerich peptides and of a novel insect defensin. J Insect Physiol 42:81–89. CrossRefGoogle Scholar
  13. Cociancich S et al (1994) Novel inducible antibacterial peptides from a hemipteran insect, the sap-sucking bug Pyrrhocoris apterus. Biochem J 300(Pt 2):567–575CrossRefGoogle Scholar
  14. Florin T et al (2017) An antimicrobial peptide that inhibits translation by trapping release factors on the ribosome. Nat Struct Mol Biol 24:752–757. CrossRefPubMedPubMedCentralGoogle Scholar
  15. Frolova L et al (1999) Mutations in the highly conserved GGQ motif of class 1 polypeptide release factors abolish the ability of human eRF1 to trigger peptidyl-tRNA hydrolysis. RNA 5:1014–1020CrossRefGoogle Scholar
  16. Gagnon MG, Roy RN, Lomakin IB, Florin T, Mankin AS, Steitz TA (2016) Structures of proline-rich peptides bound to the ribosome reveal a common mechanism of protein synthesis inhibition. Nucleic Acids Res 44:2439–2450. CrossRefPubMedPubMedCentralGoogle Scholar
  17. Gennaro R, Skerlavaj B, Romeo D (1989) Purification, composition, and activity of two bactenecins, antibacterial peptides of bovine neutrophils. Infect Immun 57:3142–3146PubMedPubMedCentralGoogle Scholar
  18. Graf M et al (2017) Proline-rich antimicrobial peptides targeting protein synthesis. Nat Prod Rep 34:702–711. CrossRefPubMedGoogle Scholar
  19. Huttner KM, Lambeth MR, Burkin HR, Burkin DJ, Broad TE (1998) Localization and genomic organization of sheep antimicrobial peptide genes. Gene 206:85–91CrossRefGoogle Scholar
  20. Knappe D et al (2010) Oncocin (VDKPPYLPRPRPPRRIYNR-NH2): a novel antibacterial peptide optimized against gram-negative human pathogens. J Med Chem 53:5240–5247. CrossRefPubMedGoogle Scholar
  21. Knappe D, Zahn M, Sauer U, Schiffer G, Strater N, Hoffmann R (2011) Rational design of oncocin derivatives with superior protease stabilities and antibacterial activities based on the high-resolution structure of the oncocin-DnaK complex. Chembiochem 12:874–876. CrossRefPubMedGoogle Scholar
  22. Knappe D, Adermann K, Hoffmann R (2015) Oncocin Onc72 is efficacious against antibiotic-susceptible Klebsiella pneumoniae ATCC 43816 in a murine thigh infection model. Biopolymers 104:707–711. CrossRefPubMedGoogle Scholar
  23. Krizsan A, Volke D, Weinert S, Strater N, Knappe D, Hoffmann R (2014) Insect-derived proline-rich antimicrobial peptides kill bacteria by inhibiting bacterial protein translation at the 70S ribosome. Angew Chem Int Ed Eng 53:12236–12239. CrossRefGoogle Scholar
  24. Krizsan A, Prahl C, Goldbach T, Knappe D, Hoffmann R (2015) Short proline-rich antimicrobial peptides inhibit either the bacterial 70S ribosome or the assembly of its large 50S subunit. Chembiochem 16:2304–2308. CrossRefPubMedGoogle Scholar
  25. Lai Y, Gallo RL (2009) AMPed up immunity: how antimicrobial peptides have multiple roles in immune defense. Trends Immunol 30:131–141. CrossRefPubMedPubMedCentralGoogle Scholar
  26. Mardirossian M, Grzela R, Giglione C, Meinnel T, Gennaro R, Mergaert P, Scocchi M (2014) The host antimicrobial peptide Bac71-35 binds to bacterial ribosomal proteins and inhibits protein synthesis. Chem Biol 21:1639–1647. CrossRefPubMedGoogle Scholar
  27. Mardirossian M et al (2018) The dolphin proline-rich antimicrobial peptide Tur1A inhibits protein synthesis by targeting the bacterial ribosome. Cell Chem Biol 25(5):530–539. CrossRefPubMedPubMedCentralGoogle Scholar
  28. Mattiuzzo M, Bandiera A, Gennaro R, Benincasa M, Pacor S, Antcheva N, Scocchi M (2007) Role of the Escherichia coli SbmA in the antimicrobial activity of proline-rich peptides. Mol Microbiol 66:151–163. CrossRefPubMedGoogle Scholar
  29. Mora L, Heurgue-Hamard V, Champ S, Ehrenberg M, Kisselev LL, Buckingham RH (2003) The essential role of the invariant GGQ motif in the function and stability in vivo of bacterial release factors RF1 and RF2. Mol Microbiol 47:267–275CrossRefGoogle Scholar
  30. Nissen P, Hansen J, Ban N, Moore PB, Steitz TA (2000) The structural basis of ribosome activity in peptide bond synthesis. Science 289:920–930CrossRefGoogle Scholar
  31. Otvos L Jr (2002) The short proline-rich antibacterial peptide family. Cell Mol Life Sci 59:1138–1150CrossRefGoogle Scholar
  32. Otvos L Jr et al (2000) Interaction between heat shock proteins and antimicrobial peptides. Biochemistry 39:14150–14159CrossRefGoogle Scholar
  33. Peschel A, Sahl HG (2006) The co-evolution of host cationic antimicrobial peptides and microbial resistance. Nat Rev Microbiol 4:529–536. CrossRefPubMedGoogle Scholar
  34. Podda E, Benincasa M, Pacor S, Micali F, Mattiuzzo M, Gennaro R, Scocchi M (2006) Dual mode of action of Bac7, a proline-rich antibacterial peptide. Biochim Biophys Acta 1760:1732–1740. CrossRefPubMedGoogle Scholar
  35. RNAcentral (2017) RNAcentral: a comprehensive database of non-coding RNA sequences. Nucleic Acids Res 45:D128–D134. CrossRefGoogle Scholar
  36. Roy RN, Lomakin IB, Gagnon MG, Steitz TA (2015) The mechanism of inhibition of protein synthesis by the proline-rich peptide oncocin. Nat Struct Mol Biol 22:466–469. CrossRefPubMedPubMedCentralGoogle Scholar
  37. Runti G et al (2013) Functional characterization of SbmA, a bacterial inner membrane transporter required for importing the antimicrobial peptide Bac7(1-35). J Bacteriol 195:5343–5351. CrossRefPubMedPubMedCentralGoogle Scholar
  38. Sato NS, Hirabayashi N, Agmon I, Yonath A, Suzuki T (2006) Comprehensive genetic selection revealed essential bases in the peptidyl-transferase center. Proc Natl Acad Sci U S A 103:15386–15391. CrossRefPubMedPubMedCentralGoogle Scholar
  39. Schnapp D, Kemp GD, Smith VJ (1996) Purification and characterization of a proline-rich antibacterial peptide, with sequence similarity to bactenecin-7, from the haemocytes of the shore crab, Carcinus maenas. Eur J Biochem 240:532–539CrossRefGoogle Scholar
  40. Schneider M, Dorn A (2001) Differential infectivity of two Pseudomonas species and the immune response in the milkweed bug, Oncopeltus fasciatus (Insecta: Hemiptera). J Invertebr Pathol 78:135–140. CrossRefPubMedGoogle Scholar
  41. Scocchi M, Lüthy C, Decarli P, Mignogna G, Christen P, Gennaro R (2009) The proline-rich antibacterial peptide Bac7 binds to and inhibits in vitro the molecular chaperone DnaK. Int J Pept Res Ther 15:147–155. CrossRefGoogle Scholar
  42. Scocchi M, Tossi A, Gennaro R (2011) Proline-rich antimicrobial peptides: converging to a non-lytic mechanism of action. Cell Mol Life Sci 68:2317–2330. CrossRefPubMedGoogle Scholar
  43. Seefeldt AC et al (2015) The proline-rich antimicrobial peptide Onc112 inhibits translation by blocking and destabilizing the initiation complex. Nat Struct Mol Biol 22:470–475. CrossRefPubMedGoogle Scholar
  44. Seefeldt AC et al (2016) Structure of the mammalian antimicrobial peptide Bac7(1-16) bound within the exit tunnel of a bacterial ribosome. Nucleic Acids Res 44:2429–2438. CrossRefPubMedPubMedCentralGoogle Scholar
  45. Seit-Nebi A, Frolova L, Justesen J, Kisselev L (2001) Class-1 translation termination factors: invariant GGQ minidomain is essential for release activity and ribosome binding but not for stop codon recognition. Nucleic Acids Res 29:3982–3987CrossRefGoogle Scholar
  46. Shamova O, Brogden KA, Zhao C, Nguyen T, Kokryakov VN, Lehrer RI (1999) Purification and properties of proline-rich antimicrobial peptides from sheep and goat leukocytes. Infect Immun 67:4106–4111PubMedPubMedCentralGoogle Scholar
  47. Shao S, Murray J, Brown A, Taunton J, Ramakrishnan V, Hegde RS (2016) Decoding mammalian ribosome-mRNA states by translational GTPase complexes. Cell 167:1229–1240.e1215. CrossRefPubMedPubMedCentralGoogle Scholar
  48. Shaw JJ, Green R (2007) Two distinct components of release factor function uncovered by nucleophile partitioning analysis. Mol Cell 28:458–467. CrossRefPubMedPubMedCentralGoogle Scholar
  49. Stensvag K, Haug T, Sperstad SV, Rekdal O, Indrevoll B, Styrvold OB (2008) Arasin 1, a proline-arginine-rich antimicrobial peptide isolated from the spider crab, Hyas araneus. Dev Comp Immunol 32:275–285. CrossRefPubMedGoogle Scholar
  50. Storici P, Zanetti M (1993) A cDNA derived from pig bone marrow cells predicts a sequence identical to the intestinal antibacterial peptide PR-39. Biochem Biophys Res Commun 196:1058–1065. CrossRefPubMedGoogle Scholar
  51. Thompson J et al (2001) Analysis of mutations at residues A2451 and G2447 of 23S rRNA in the peptidyltransferase active site of the 50S ribosomal subunit. Proc Natl Acad Sci U S A 98:9002–9007CrossRefGoogle Scholar
  52. Voss NR, Gerstein M, Steitz TA, Moore PB (2006) The geometry of the ribosomal polypeptide exit tunnel. J Mol Biol 360:893–906CrossRefGoogle Scholar
  53. Yeaman MR, Yount NY (2003) Mechanisms of antimicrobial peptide action and resistance. Pharmacol Rev 55:27–55. CrossRefPubMedGoogle Scholar
  54. Zahn M, Berthold N, Kieslich B, Knappe D, Hoffmann R, Strater N (2013) Structural studies on the forward and reverse binding modes of peptides to the chaperone DnaK. J Mol Biol 425:2463–2479. CrossRefPubMedGoogle Scholar
  55. Zahn M, Kieslich B, Berthold N, Knappe D, Hoffmann R, Strater N (2014) Structural identification of DnaK binding sites within bovine and sheep bactenecin Bac7. Protein Pept Lett 21:407–412CrossRefGoogle Scholar
  56. Zanetti M, Litteri L, Gennaro R, Horstmann H, Romeo D (1990) Bactenecins, defense polypeptides of bovine neutrophils, are generated from precursor molecules stored in the large granules. J Cell Biol 111:1363–1371CrossRefGoogle Scholar
  57. Zanetti M, Litteri L, Griffiths G, Gennaro R, Romeo D (1991) Stimulus-induced maturation of probactenecins, precursors of neutrophil antimicrobial polypeptides. J Immunol 146:4295–4300PubMedGoogle Scholar
  58. Zanetti M, Del Sal G, Storici P, Schneider C, Romeo D (1993) The cDNA of the neutrophil antibiotic Bac5 predicts a pro-sequence homologous to a cysteine proteinase inhibitor that is common to other neutrophil antibiotics. J Biol Chem 268:522–526PubMedGoogle Scholar
  59. Zasloff M (2002) Antimicrobial peptides in health and disease. N Engl J Med 347:1199–1200. CrossRefPubMedGoogle Scholar
  60. Zavialov AV, Mora L, Buckingham RH, Ehrenberg M (2002) Release of peptide promoted by the GGQ motif of class 1 release factors regulates the GTPase activity of RF3. Mol Cell 10:789–798CrossRefGoogle Scholar

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© Springer Nature Singapore Pte Ltd. 2019

Authors and Affiliations

  1. 1.Institute for Biochemistry and Molecular BiologyUniversity of HamburgHamburgGermany

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