kalata B1(1) Protein Card

General Information
Name kalata B1
Class Cyclotide
Technique MS,EST,PCR,NGS,Amino acid analysis
Average Mass 2892.28
Monoisotopic Mass 2890.14
m/z M+H 2892.28
ProteinType Wild type
Organism Viola odorata
Oldenlandia affinis
Viola tricolor
Hybanthus enneaspermus
Viola baoshanensis
Viola yedoensis
Viola philippica
Viola sumatrana Miq
Notes <p>Hellinger et al. identified a possible deamination of Asparagine or Glutamine residues and a possible tryptophan oxidation to 3-hydroxy-tryptophan.</p> <p>Was found in transcriptome data of Hybanthus enneaspermus (Du et al., 2020).</p>
Cyclic Yes

Chemical Shift
Chemical shift data for kalata B1 [...]
[...]Niyomploy et al. (2016) Peptide Science 106:796-805
The cyclotides caused a concentration-dependent response in the LDA with the two nematode species. [...]Colgrave ML et al. (2008) Biochemistry 47:5581-5589
Motility affected by cyclotides [...]Colgrave ML et al. (2008) Chembiochem 9:1939-1945
kB1 and kB2 inhibited the hatching of H. contortus and T. colubriformis eggs in a concentration-dependent manner. [...]Colgrave ML et al. (2008) Biochemistry 47:5581-5589
A range of natural variants were selected based on net charge and/or hydrophobicity and were tested alongside the prototypic cyclotide kB1 in larval development assays with H. contortus and T. colubriformis. [...]Colgrave ML et al. (2008) Chembiochem 9:1939-1945
A range of concentrations of kB1 was tested and scoring was conducted after 24, 48 and 72 h. After 24 h, motility scores of 0 were recorded at the two highest concentrations tested and the intermediate concentrations responded in a dose-dependent manner. After 48 h, there was a significant reduction in motility and, after 72 h, motility scores of 0 were recorded for all concentrations tested except the lowest two (<25 µg/mL). It was also noted that while in the control assay tubes, a large proportion of laid eggs hatched to L1 larvae after 24 h, very few eggs hatched in the kB1 treated assay tubes above 400 µg/mL and nearly all newly hatched larvae were dead above kB1 concentrations of 50 µg/mL. [...]Colgrave ML et al. (2008) Biochemistry 47:5581-5589
Anti-HIV activity shown for kalata B1 but not for acyclic permutants. [...]Daly NL et al. (2004) FEBS Lett 574:69-72
All cyclotides tested from V. yedoensis demonstrated anti-HIV activity, with cycloviolacin Y5 being the most potent. [...]Wang CK et al. (2007) J Nat Prod 71:47-52
Tricyclon A has minimal haemolytic activity [...]Mulvenna JP et al. (2005) Structure (Camb) 13:691-701
Substitution of certain residues of kalata B1 to alanine result in loss of hemolytic activity. Residues in loops 1 and 3 along with a few other isolated residues strongly affect activity. [...]Simonsen SM et al. (2008) J Biol Chem 283:9805-9813
Hemolytic activity for established for kalata B1 but not acyclic permutants [...]Daly NL and Craik DJ (2000) J Biol Chem 275:19068-75
Hemolytic activity established for kalata B1, circulin A, circulin B and cyclopsychotride A [...]Tam JP et al. (1999) Proc Natl Acad Sci U S A 96:8913-8
Cycloviolacins Y4 and Y5 are the most hemolytic of the new cyclotides, both being more potent than the prototypic cyclotide kalata B1. By contrast, cycloviolacin Y1 is substantially less active than all of the other cyclotides tested. [...]Wang CK et al. (2007) J Nat Prod 71:47-52
Cycloviolacin H4 has potent haemolytic activity compared to kalata B1 [...]Chen B et al. (2006) J Nat Prod 69:23-8
Both [P20D/V21K]kalata B1 and [W19K/P20N/V21K]kalata B1 exhibited no haemolytic activity. Kalata B1 exhibited mild haemolytic activity. [...]Clark RJ et al. (2006) Biochem J 394:85-93
Both kalata B1 oia and B1 nfk exhibited no haemolytic activity [...]Plan MR et al. (2007) Chembiochem 8:1001-1011
At a concentration of 25 ºM, a more than 6-fold difference exists between the most hemolyticcyclotide, cycloviolacin O24 (~75% hemolysis), and the least hemolytic cyclotide cycloviolacin O14 (~11% hemolysis). The sensitivity of hemolytic activity to variations in the peptide sequence is evident when comparing cycloviolacin O2 and O13. Here, the only sequence deviation is a single residue substitution of a serine in loop 3 of O2 (HD50 ~36 ºM) to an alanine in the homologous position in O13 (HD50 ~11 ºM). The loss of this single hydroxy group changes the HD50 more than three-fold. [...]Ireland DC et al. (2006) Biochem J. 400:1-12
Cyclotides show molluscicidal activity [...]Plan MR et al. (2008) J Agric Food Chem 56:5237-5241
Enzymatic Digest
Resistance to enzymatic digest shown for kalata B1 and acyclic permutants. Reduced protein was susceptible. [...]Colgrave ML and Craik DJ (2004) Biochemistry 43:5965-75
G6A and E7A mutants of kalata B1 were completely impervious to enzymatic digestion by the three endoproteinases trypsin, chymotrypsin, and proteinase K, as was the prototypic cyclotide kalata B1. [...]Simonsen SM et al. (2008) J Biol Chem 283:9805-9813
Substitution of certain residues of kalata B1 to alanine result in loss of insecticidal activity [...]Simonsen SM et al. (2008) J Biol Chem 283:9805-9813
Kalata B1 shown to possess potent insecticidal activity. [...]Jennings C et al. (2001) Proc Natl Acad Sci U S A 98:10614-9
kalata B1 induces disruption of the microvilli, blebbing, swelling, and ultimately rupture of the cells of the gut epithelium [...]Barbeta BL et al. (2008) Proc Natl Acad Sci U S A 105:1221-1225
H. armigera larvae fed kalata B1 or kalata B2 exhibited reduced growth and higher mortality rate compared to cyclotide-less diet [...]Jennings CV et al. (2005) Biochemistry 44:851-60
Growth of H. armigera larvae stunted by kalata B1 containing diet [...]Barbeta BL et al. (2008) Proc Natl Acad Sci U S A 105:1221-1225
Metal binding assay
Suggests that kalata B1 may contain a metal binding site. [...]Skjeldal L et al. (2002) Arch Biochem Biophys 399:142-8
Kalata B1 binds to Mn2+ in the presence of DPC micelles [...]Shenkarev ZO et al. (2006) FEBS J 273:2658-2672
In the presence of Mn2+ kalata B1, T16A and T27A showed specific broadening of a defined set of amide signals. There was almost complete broadening of the amide signals from C14, N15, and T16/A16, and significant broadening (i.e. ~70% attenuation) of loop 1 residues. By contrast, E7A showed significantly less broadening. [...]Simonsen SM et al. (2008) J Biol Chem 283:9805-9813
Chemical stability
Kalata B1 was found to be resistant to both chemical and thermal denaturation. [...]Colgrave ML and Craik DJ (2004) Biochemistry 43:5965-75
Kalata B1 and B2 and acyclic permutants were found to be resistant to chemical denaturation in the oxidised state but not when reduced. [...]Colgrave ML and Craik DJ (2004) Biochemistry 43:5965-75
Membrane Binding Assay
Showed that kalata B1 and B6 would bind selectively to bacterial membranes. [...]Kamimori H et al. (2005) Anal Biochem 337:149-53
Kalata B1 binds to DPC micelles. [...]Shenkarev ZO et al. (2006) FEBS J 273:2658-2672
Kalata B1 and B2 cause leakage of dye-containing POPC vesicles. [...]Colgrave ML et al. (2008) Biochemistry 47:5581-5589
Protease inhibition
Kalata B1 and B2 were found to have no trypsin or chymotrypsin inhibitory activity. [...]Jennings C et al. (2001) Proc Natl Acad Sci U S A 98:10614-9
Acid Hydrolysis
Kalata B1 was observed to be resistant to acid hydrolysis. [...]Colgrave ML and Craik DJ (2004) Biochemistry 43:5965-75
Kalata B1, circulin A and B and cyclopsychotride shown to possess some anti-bacterial activity. [...]Tam JP et al. (1999) Proc Natl Acad Sci U S A 96:8913-8
Reductive Unfolding
Lowering the pH to 4.5 and using TCEP (5-300 mM) as the reductant revealed the presence of minor components along the unfolding pathway of kalata B1. [...]Daly NL et al. (2003) J Biol Chem 278:6314-22
NOE Restraint
NOE restraints for 1NB1 [...]
NOE restraints for 1KAL [...]
Cancer Cell Toxicity
CyO2, CyO13, kalata B1, and varv peptide A exhibited dose-dependent cytotoxicity in MTT assays with IC50 values of 2.15-7.93 uM against human brain astrocytoma cells (U-87 MG) and human bone marrow derived neuroblastoma cells (SH-SY5Y). CyO2 and varv peptide A also sensitized SH-SY5Y and U-87 MG cells to temozolomide (TMZ) which is the only effective chemotherapeutic drug used to treat glioblastoma multiforme (GBM). [...]Gerlach,S.L. et al. (2022) J Nat Prod 85:34-46

Jennings CV, Rosengren KJ, Daly NL, Plan M, Stevens J, Scanlon MJ, Waine C, Norman DG, Anderson MA, Craik DJ (2005) Isolation, solution structure, and insecticidal activity of kalata B2, a circular protein with a twist: do Mobius strips exist in nature? Biochemistry 44:851-60
Daly NL, Gustafson KR, Craik DJ (2004) The role of the cyclic peptide backbone in the anti-HIV activity of the cyclotide kalata B1. FEBS Lett 574:69-72
Simonsen SM, Daly NL, Craik DJ (2004) Capped acyclic permutants of the circular protein kalata B1. FEBS Lett 577:399-402
Dutton JL, Renda RF, Waine C, Clark RJ, Daly NL, Jennings CV, Anderson MA, Craik DJ (2004) Conserved structural and sequence elements implicated in the processing of gene-encoded circular proteins. J Biol Chem 279:46858-67
Colgrave ML, Craik DJ (2004) Thermal, chemical, and enzymatic stability of the cyclotide kalata B1: the importance of the cyclic cystine knot. Biochemistry 43:5965-75
Goransson U, Craik DJ (2003) Disulfide mapping of the cyclotide kalata B1. Chemical proof of the cystic cystine knot motif. J Biol Chem 278:48188-96
Barry DG, Daly NL, Clark RJ, Sando L, Craik DJ (2003) Linearization of a naturally occurring circular protein maintains structure but eliminates hemolytic activity. Biochemistry 42:6688-95
Daly NL, Clark RJ, Craik DJ (2003) Disulfide folding pathways of cystine knot proteins. Tying the knot within the circular backbone of the cyclotides. J Biol Chem 278:6314-22
Jennings C, West J, Waine C, Craik D, Anderson M (2001) Biosynthesis and insecticidal properties of plant cyclotides: the cyclic knotted proteins from Oldenlandia affinis. Proc Natl Acad Sci U S A 98:10614-9
Daly NL, Craik DJ (2000) Acyclic permutants of naturally occurring cyclic proteins. Characterization of cystine knot and beta-sheet formation in the macrocyclic polypeptide kalata B1. J Biol Chem 275:19068-75
Tam JP, Lu YA, Yang JL, Chiu KW (1999) An unusual structural motif of antimicrobial peptides containing end-to-end macrocycle and cystine-knot disulfides. Proc Natl Acad Sci U S A 96:8913-8
Kamimori H, Hall K, Craik DJ, Aguilar MI (2005) Studies on the membrane interactions of the cyclotides kalata B1 and kalata B6 on model membrane systems by surface plasmon resonance. Anal Biochem 337:149-53
Nourse A, Trabi M, Daly NL, Craik DJ (2004) A comparison of the self-association behavior of the plant cyclotides kalata B1 and kalata B2 via analytical ultracentrifugation. J Biol Chem 279:562-70
Rosengren KJ, Daly NL, Plan MR, Waine C, Craik DJ (2003) Twists, knots, and rings in proteins. Structural definition of the cyclotide framework. J Biol Chem 278:8606-16
Skjeldal L, Gran L, Sletten K, Volkman BF (2002) Refined structure and metal binding site of the kalata B1 peptide. Arch Biochem Biophys 399:142-8
Daly NL, Love S, Alewood PF, Craik DJ (1999) Chemical synthesis and folding pathways of large cyclic polypeptides: studies of the cystine knot polypeptide kalata B1. Biochemistry 38:10606-14
Saether O, Craik DJ, Campbell ID, Sletten K, Juul J, Norman DG (1995) Elucidation of the primary and three-dimensional structure of the uterotonic polypeptide kalata B1. Biochemistry 34:4147-58
Ireland DC, Colgrave ML, Craik DJ (2006) A novel suite of cyclotides from Viola odorata: sequence variation and the implications for structure, function and stability. Biochem J. 400:1-12
Plan MR, Goransson U, Clark RJ, Daly NL, Colgrave ML, Craik DJ (2007) The Cyclotide Fingerprint in Oldenlandia affinis: Elucidation of Chemically Modified, Linear and Novel Macrocyclic Peptides Chembiochem 8:1001-1011
Wang CK, Colgrave ML, Gustafson KR, Ireland DC, Göransson U, Craik DJ (2007) Anti-HIV Cyclotides from the CHinese Medicinal Herb Viola yedoensis J Nat Prod 71:47-52
Barbeta BL, Marshall AT, Gillon AD, Craik DJ, Anderson MA (2008) Plant cyclotides disrupt epithelial cells in the midgut of lepidopteran larvae Proc Natl Acad Sci U S A 105:1221-1225
Qin Q, McCallum EJ, Kaas Q, Suda J, Saska I, Craik DJ, Mylne JS. (2010) Identification of candidates for cyclotide biosynthesis and cyclisation by expressed sequence tag analysis of Oldenlandia affinis. BMC Genomics 11:111-0
He,W., Chan,L.Y., Zeng,G., Daly,N.L., Craik,D.J. and Tan,N. (2011) Isolation and characterization of cytotoxic cyclotides from Viola philippica. Peptides 32:1719-1723
Sando,L., Henriques,S.T., Foley,F., Simonsen,S.M., Daly,N.L., Hall,K.N., Gustafson,K.R., Aguilar,M.I. and Craik,D.J. (2011) A synthetic mirror image of kalata B1 reveals that cyclotide activity is independent of a protein receptor. Chembiochem 12:2456-2462
Cascales,L., Henriques,S.T., Kerr,M.C., Huang,Y.H., Sweet,M.J., Daly,N.L. and Craik,D.J. (2011) Identification and characterization of a new family of cell-penetrating peptides: cyclic cell-penetrating peptides. J. Biol. Chem. 286:36932-36943
Burman,R., Strömstedt,A.A., Malmsten,M. and Göransson,U. (2011) Cyclotide-membrane interactions: defining factors of membrane binding, depletion and disruption. Biochim. Biophys. Acta 1808:2665-2673
Hellinger,R., Koehbach,J., Soltis,D.E., Carpenter,E.J., Wong,G.K. and Gruber,C.W. (2015) Peptidomics of Circular Cysteine-Rich Plant Peptides: Analysis of the Diversity of Cyclotides from Viola tricolor by Transcriptome and Proteome Mining. J. Proteome Res. 14:4851-4862
Poth, A.G., Huang, Y.H., Le, T.T., Kan, M.W. and Craik, D.J., (2019) Pharmacokinetic characterization of kalata B1 and related therapeutics built on the cyclotide scaffold Int J Pharm 565:437-446
Niyomploy, P., Chan, L.Y., Poth, A.G., Colgrave, M.L., Sangvanich, P. and Craik, D.J. (2016) Discovery, isolation, and structural characterization of cyclotides from Viola sumatrana Miq Peptide Science 106:796-805
Du,Q., Chan,L.Y., Gilding,E.K., Henriques,S.T., Condon,N.D., Ravipati,A.S., Kaas,Q., Huang,Y.H. and Craik,D.J. (2020) Discovery and mechanistic studies of cytotoxic cyclotides from the medicinal herb Hybanthus enneaspermus. J Biol Chem 295:10911-10925
Aslam,L., Kaur,R., Sharma,V., Kapoor,N. and Mahajan,R. (2021) Isolation and characterization of cyclotides from the leaves of Viola odorata L. using peptidomic and bioinformatic approach. 3 Biotech 11:211-0
Gerlach,S.L., Dunlop,R.A., Metcalf,J.S., Banack,S.A., and Cox,P.A. (2022) Cyclotides Chemosensitize Glioblastoma Cells to Temozolomide. J Nat Prod 85:34-46

Protein precursor(s) cyclotide precursor 6b
cyclotide precursor 7c
kalata B1 precursor
cyclotide k1 precursor
Oak10 precursor
kalata B1 precursor
Nucleic acids kalata B1 precursor [OaK1]
cyclotide k1 precursor
cyclotide precursor 6b mRNA
cyclotide precursor 7c mRNA
Structure High-resolution solution structure of kalata B1
Refined structure and disulfide pairing of the kalata B1 peptide
kalata B1
Structure of cyclotide Kalata B1 in DPC micelles solution
Refined structure and disulfide pairing of the kalata B1 peptide
Solution structure of the all-D kalata B1
Racemic structure of kalata B1 (kB1)
Links GenBank AAL05477.1
SwissProt P56254