Professor John Bowie

Research Interests

1. Biologically-active host-defence peptides and pheromones
   

Recent work in this area has involved an immunomodulator peptide isolated from a wallaby, a number of defence peptides, a male sex pheromone from an Australian tree frog and an nNOS active peptide from a South American spider. In particular:

(a) The lactating female of the Tammar wallaby (Macropus eugenii) produces a potent peptide (eugenin) which enhances killer T cell formation in her pouch protecting the young from bacterial flora and other predators [1]. Similar activity is found in disulfide containing peptides from the skin secretions of a number of froglets of the genus Crinia, e.g. riparin 1 [2].

Eugenin       pEQDY(SO₃H)VFMHPF-NH₂
Riparin 1      RLCIPVIFPC-OH
Caerin 1.8    GLFKVLGSVAKHLLPHVVPVIAEKL-NH₂

(b) Australian frogs contain, in their skin glands, membrane active antibiotic peptides like caerin 1.8 [2] and fallaxidin 4.1 [3]. Nuclear magnetic resonance and associated techniques are used to determine the 3D structure of these peptides and the mechanisms by which they penetrate (and destroy) bacterial cell membranes [3,4].

Fallaxidin 4.1  GLLSFLPKVIGVIGHLIHPPS-OH

(c) As well as antibiotics and neuropeptides, Australian frogs produce other active peptides  which, when secreted onto their skin inhibit the production of the chemical messenger nitric oxide by the nitric oxide synthase isoforms. Ingestion of these peptides may either incapacitate or kill a predator. The peptides act by complexing with the regulatory peptide (molecular switch) Ca²⁺ calmodulin which is then unable to attach to the calmodulin docking domain of the appropriate nitric oxide synthase [5]. The helix-hinge-helix peptide caerin 1.8, is one of the most potent of this class of peptide. This work has been reviewed [2,6] and is ongoing.

(d) cDNA sequencing has shown that antibiotic peptides from the genus Litoria originated from the same ancestor gene from which many northern hemisphere amphibian peptides were formed some 150 million years ago [2,cf. 7].  In contrast, disulfide containing peptides like riparin 1 from froglets of the Crinia genus may have been formed following the separation of Gondwana to form Australia some 90 million years ago [8].

Splendipherin  GLVSSIGKALGGLLADVVKSKGQPA-OH

(e) An ongoing interest is the identification and characterisation of aqueous pheromones.  The first male sex pheromone of a tree frog was identified as splendipherin, the male sex pheromone of Litoria splendida [9,10].  This peptide has been shown to move across the surface of water at a concentration of less than nanomolar [11].  Our interest in this area is continuing and we have been searching for the sex pheromone of the Cane Toad (Bufo marinus) for some years.

(f) We are currently investigating, together with Prof. John Carver and Dr Tara Pukala, those peptides (that we have isolated from the skin glands of frogs and toads) which inhibit the aggregation of those peptides which are associated with Alzheimer's and Parkinson's diseases.  These studies have also uncovered a number of amphibian antibiotic peptides which self aggregate to form fibrils.

(g) We are also studying how the replacement of Asp with isoAsp in selected neuropeptides and membrane active peptides changes the shape, activity and cytotoxicity of those peptides.  To date we have demonstrated that initiating this change to Asp3 of uperin 1.1 significantly enhances smooth muscle activity, making this the most active  of the physalaemin family of neuropeptides (Calabrese and Markulic unpublished).  This work is ongoing.

Uperin 1.1    pEADPNAFYGLM-NH2

(h) The venom of the South American spider Cupiennius salei, contains four 35 residue cupiennin peptides. These peptides, e.g. cupiennin 1a, are known antimicrobial agents, but it has now shown that they are more potent in deactivating nNOS than the amphibian peptides described above, and that they also operate by complexing with calmodulin. This may assist with paralysis of the prey following envenomation [12,13].

Cupiennin 1a   GFGALFKFLAKKVAKTVAKQAAKQGAKYVVNKQME-NH₂

2. Ion chemistry (mass spectrometry)

A major research interest over the years has been the study of negative ion mass spectrometry. Our recent work in this area is currently involved in the use of negative ions to (a) to form neutrals of stellar significance [14,15], (b) develop the use of negative ion mass spectrometry as a viable analytical method for the sequence determination of peptides and proteins [16,17], and (c) to use ion-molecule chemistry of transient anions to study the structure and reactivity of these anions in the gas phase [18].

(a) One of the most important questions in science is how did life originate on our planet. Aristotle [19]considered that the five "elements" were extraterrestrial, and later Hoyle, [20] proposed  that the primary chemicals of life originate in space, i.e. by reactions in interstellar dust clouds, circumstellar envelopes or interstellar ice. We strip an electron from a negative ion in the mass spectrometer to produce a reactive neutral of known bond connectivity which has been identified in interstellar media. We study the structure and reactivity of this neutral in the mass spectrometer in order to assess the likelihood that it is long-lived enough to be able to undergo reaction to form, for example, an amino acid, a sugar or a nucleotide base. As an example, we have shown that electron stripping of ^-CH₂CN produces the stable radical ^.CH₂CN which may then produce the glycine precursor NH₂CH₂CN which can then be hydrolysed in interstellar ice (around comets, planets etc) to yield the amino acid glycine [21]. We are currently considering ways by which other amino acids can be formed in stellar regions. We have reported that the pyrimidine nucleobase uracil may be produced by the reaction between CCCO and urea and that this reaction is more kinetically favourable if the urea is monohydrated [22].

^.CH₂CN + NH_2^. → NH₂CH₂CN → NH₂CH₂CO₂H (glycine)

The prototypical two carbon sugar glycolaldehyde (HOCH2CHO) has been detected towards the galactic centre source Sagittarius B2(N) by means of rotational transitions [23].  We are currently studying, by experiment and theory in concert, the possible interstellar synthetic pathways to form glycolaldehyde and higher sugars [24].

We are also interested in cumulenes that have been detected in interstellar molecular clouds and are currently studying linear CCCX and their planar rhombic isomers. These are formed by the charge stripping of negative ions in a mass spectrometer, e.g. CCCC [25,26] and CCCN [27].  Cumulenes like CCCB [28] and NCCCCCN [29] have also been studied. This experimental work is complemented by high-level theoretical studies (computational chemistry) in order to probe the structure and reactivity of these transient cumulenes [30].

(b) Positive ion mass spectrometry is the normal way to sequence peptides and proteins.  We have been exploring the use of negative ion electrospray mass spectrometry for the sequencing of peptides, and this work has been reviewed [16,17].  We have recently shown that negative ion mass spectrometry is the method of choice for the identification of post-translational modifications including disulfides  [31].  We are currently investigating the use of negative ions to identify and position disulfide functionality in disulfide containing proteins [32], and the rearrangement processes of the (M-H)- anions of phosphorylated peptides [33].

(c) Finally, we are currently using ion molecule chemistry in a modified ion-trap mass spectrometer to investigate the structures of transient carbanions in the gas phase; e.g. the reaction between a carbanion and carbon disulfide to form an adduct whose subsequent fragmentations provide information concerning the structure of the original carbanion.  This work is also complemented by theoretical studies [18].

References

[1]  R.V.Baudinette, P.Boontheung, I.F.Musgrave, P.A.Wabnitz, V.M.Maselli, J.Skinner, P.F.Alewood, C.S.Brinkworth and J.H.Bowie, FEBS J., 2005, 272, 433-443.

[2]  T.L.Pukala, J.H.Bowie, V.M.Maselli, I.F.Musgrave and M.J.Tyler, Nat. Prod. Rep., 2006, 23, 368-393.

[3]  P.J.Sherman, R.J.Jackway, J.D.Gehman, S.Praporski, G.A.McCubbin, A.Mechler, L.L.Martin, F.Separovic and J.H.Bowie, Biochemistry 2009, 48, 11892-11901.

[4]  G.A.McCubbin, S.Praporski, S.Piantavigna, J.H.Bowie, F.Separovic, R.Hoffmann and L.L.Martin, Eur. Biophys. J., 2011, 40, 437-446.

[5]  J.R.Doyle, C.S.Brinkworth, K.L.Wegener, J.A.Carver, L.E.Llewellyn, I.N.Olver, J.H.Bowie, P.A.Wabnitz and M.J.Tyler, Eur. J. Biochem., 2003, 270, 1141-1153.

[6]   J.R.Doyle, J.H.Bowie, R.J.Jackway, L.E.Llewellyn, T.L.Pukala, M.A.Apponyi and G.W.Booker, Anuran host-defence peptides that complex with calcium calmodulin and inhibit the synthesis of the cell signaling agent nitric oxide by neuronal nitric oxide synthase., in Bioactive Peptides, eds, J.Howl and S.Jones, CRC Press, 2009, chapter 14, pp. 315-333.

[7]  D.Vanhoye, F.Bruston, P.Nicolas and M.Amiche, Eur. J. Biochem., 2003, 270, 2068-2081.

[8]  R.J.Jackway, T.L.Pukala, V.M.Maselli, I.F.Musgrave, J.H.Bowie, Y.Liu, K.H.Surinya-Johnson, S.C.Donnellan, J.R.Doyle, L.E,Llewellyn and M.J.Tyler.  Regul. Pept., 2008, 151, 80-87.

[9]  P.A.Wabnitz, J.H.Bowie, M.J.Tyler, J.C.Wallace and B.P.Smith, Nature 1999, 401, 444-445.

[10]  J.H.Bowie. Splendipherin: the aquatic male sex pheromone of the Magnificent Tree Frog Litoria splendida,  in Pheromones, Theories, Types and Uses.  F.Columbus ed., Nova Publishers, New York, USA, 2010, Chapter 1., pp 201-214.

[11]  A.W.Perriman, M.A.Apponyi, M.A.Buntine, R.J.Jackway, M.W.Rutland, J.W.White and J.H.Bowie,  FEBS. J., 2008, 275, 3362-3374.

[12]  T.L.Pukala, J.R.Doyle, L.E.Llewellyn, L.Kuhn-Nerwig, F.Separovic and J.H.Bowie. FEBS J., 2007, 274, 1778-1784.

[13]   T.L.Pukala, M.P.Boland, J.D.Gehman, L.Kuhn-Nerwig, F.Separovic and J.H.Bowie. Biochemistry, 2007, 46, 3576-3585.

[14]  S.J.Blanksby and J.H.Bowie, Mass Spectrom. Rev. 1999, 18, 131-151.

[15]  S.Dua, S.J.Blanksby, S.Peppe, A.M.McAnoy and J.H.Bowie, Curr. Org. Chem., 2003, 7, 1545-1564.

[16]  J.H.Bowie, C.S.Brinkworth and S.Dua, Mass Spectrom. Rev., 2002, 21, 87-107.

[17]  J.H.Bowie and D.Bilusich, Mass Spectrom. Rev., 2009, 28, 20-34.

[18]  M.J.Maclean, S.Walker, T.Wang, P.C.H.Eichinger, P.J.Sherman and J.H.Bowie, Org. Biomol. Chem., 2010, 8, 371-377.

[19]  G.E.R.Lloyd,  Aristotle.  The Growth and Structure of his Thought.  Cambridge Univ. Press, Cambridge, UK. 1968, pp 133-139.

[20]   E.M.Burbidge, G.R. Burbidge, W.A. Fowler and F. Hoyle. Revs. Mod. Physics 1957, 29, 547-650.

[21]  H.J.Andreazza, M.Fitzgerald and J.H.Bowie, Org. Biomol. Chem., 2006, 4, 2466-2472.

[22]  T. Wang and J.H.Bowie, Org. Biomol. Chem., 2011, DOI: 10.1039/c1ob06352a

[23]   J.M.Hollis, F.J.Lovas and P.R.Jewell, Astrophys. J., 2000, 540, L107.

[24]   T.Wang and J.H.Bowie, Org. Biomol. Chem., 2010, 8, 4756-4766.

[25]  S.J.Blanksby, D.Schröder, S.Dua, J.H.Bowie and H.Schwarz, J. Am. Chem. Soc., 2000, 122, 7105-7113.

[26]  T.Wang, M.A.Buntine and J.H.Bowie, J. Phys. Chem. A, 2009, 113, 12952-12960.

[27]  M.J.Maclean, M.Fitzgerald and J.H.Bowie. J. Phys. Chem. A, 2007, 111, 12932-12937.

[28]  T.Wang and J.H.Bowie, Phys. Chem. Chem. Phys., 2009, 23, 7553-7561.

[29]  S.Dua, T.Wang and J.H.Bowie, J. Phys. Chem. A, 2010, 114, 949-955.

[30]  T.Wang and J.H.Bowie, Mass Spectrom. Rev., 2011, 30, 1225-1241.

[31]  D.Bilusich and J.H.Bowie,  Rapid Commun. Mass Spectrom., 2007, 21, 619-628.

[32]  H.J.Andreazza and J.H.Bowie, Phys. Chem. Chem. Phys., 2010, 12, 13400-13407.

[33]  T.Wang, D.Bilusich, H.J.Andreazza and J.H.Bowie, Rapid Commun. Mass Spectrom., 2009, 23, 1669-1677.

 

Research Awards, Biographical and other notes

Rennie Medal (Royal Australian Chemical Institute, 1967); H.G.Smith Medal (RACI, 1974); Morrison Medal (Australian and New Zealand Society of Mass Spectrometry, 1997); A.J.Birch Medal (RACI, 2001); Verco Medal (Royal Society of South Australia, 2003); J.J.Thomson Gold Medal (International Council for Mass Spectrometry, 2006); ANZSMS Medallist (2011); Org. Mass Spectrom., 1975, 10, v-vi;  Current Contents, 1981, 21, 5;  Austral. J. Chem., 2003, 56, 343-348; Encyclopaedia of Mass Spectrometry, Vol. 9, 2008; Bowie Medal of ANZSMS introduced in 2009.

Expertise for Media Contact

Categories	Science and technology
Expertise	Mass spectrometry Interstellar molecules Bioactive peptides from Australian animals
Notes	Alt phone: (08) 8303 5996

Entry last updated: Sunday, 5 Feb 2012