Presentation of scientific reports

This thesis is presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy at the College of Business, RMIT University. It includes five scientific papers which have been submitted and are currently under review or accepted for publication in targeted journals. These papers comprise one conceptual paper and four empirical papers.

259p runthenight04 02-02-2023 7 3   Download

The K103N substitution is a frequently observed HIV-1 RT mutation in patients who do not respond to combinationtherapy. The drugs Efavirenz, MSC194 and PNU142721 belong to the recent generation of NNRTIs characterized by an improved resistance profile to the most common single point mutations within HIV-1 RT, including the K103N mutation. In the present study we present structural observations from Efavirenz in complex with wild-type protein and the K103N mutant and PNU142721 and MSC194 in complex with the K103N mutant.

8p system191 01-06-2013 41 4   Download

The dinitrogenase component proteins of the conventional Mo nitrogenase (MoFe protein) and of the alternative Fe-only nitrogenase (FeFe protein) were both isolated and purified from Rhodobacter capsulatus, redox-titrated according to the same procedures and subjected to an EPR spectroscopic comparison. In the course of an oxidative titration of the MoFe protein (Rc1Mo) three significant S ¼ 1/2 EPR signals deriving from oxidized states of the P-cluster were detected: (1) a rhombic signal (g ¼ 2.07, 1.96 and 1.

12p system191 01-06-2013 31 4   Download

Mer belongs to the Tyro 3 family of receptor tyrosine kinases (RTKs). Together with Axl and Rse, the three RTKs are believed to play important functional roles in the male gonads because gene knockout male mice lacking all of these receptors are infertile. In the present study, postnatal expression of Axl and Rse in mouse testes decreased during maturation while expression of Mer increased age-dependently during testicular development.

12p system191 01-06-2013 43 2   Download

This review describes studies of particular enzymatically catalyzed reactions to investigate the possibility that catalysis is mediated by protein dynamics. That is, evolution has crafted the protein backbone of the enzyme to direct vibrations in such a fashion to speed reaction. The review presents the theoretical approach we have used to investigate this problem, but it is designed for the nonspecialist.

10p system191 01-06-2013 37 4   Download

Aralar1 and citrin are members of the subfamily of calciumbinding mitochondrial carriers and correspond to two isoforms of the mitochondrial aspartate/glutamate carrier (AGC). These proteins are activated by Ca2+ acting on the external side of the inner mitochondrial membrane. Although it is known that aralar1 is expressed mainly in skeletal muscle, heart and brain, whereas citrin is present in liver, kidney and heart, the precise tissue distribution of the two proteins in embryonic and adult tissues is largely unknown....

8p system191 01-06-2013 35 4   Download

The main component of cerebral amyloid angiopathy (CAA) in Alzheimer’s disease is the amyloid-b protein (Ab), a 4-kDa polypeptide derived from the b-amyloid protein precursor (APP). The accumulation of Ab in the basement membrane has been implicated in the degeneration of adjacent vascular smooth muscle cells (VSMC). However, the mechanism of Ab toxicity is still unclear. In this study, we examined the effect of substrate-bound Ab on VSMC in culture. The use of substrate-bound proteins in cell culture mimics presentation of the proteins to cells as if bound to the basement membrane.

9p system191 01-06-2013 43 5   Download

A clear difference in the enthalpy changes derived from spectroscopic and calorimetric measurements has recently been shown. The exact interpretation of this deviation varied from study to study, but it was generally attributed to the non-two-state transition and heat capacity change. Although the temperature-dependent thermodynamics of the duplex formation was often implied, systemic and extensive studies have been lacking in universally assigning the appropriate thermodynamic parameter sets.

10p system191 01-06-2013 37 4   Download

Trypanosoma brucei is the cause of the diseases known as sleeping sickness in humans (T. brucei ssp. gambiense and ssp. rhodesiense) and ngana in domestic animals (T. brucei brucei) in Africa. Procyclic trypomastigotes, the tsetse vector stage, express a surface-bound trans-sialidase that transfers sialic acid to the glycosylphosphatidylinositol anchor of procyclin, a surface glycoprotein covering the parasite surface. Trans-sialidase is a unique enzyme expressed by a few trypanosomatids that allows them to scavenge sialic acid from sialylated compounds present in the infected host. ...

10p system191 01-06-2013 44 4   Download

Human glandular kallikrein 2 (hK2) is a trypsin-like serine protease expressed predominantly in the prostate epithelium. Recently, hK2 has proven to be a useful marker that can be used in combination with prostate specific antigen for screening and diagnosis of prostate cancer. The cleavage by hK2 of certain substrates in the proteolytic cascade suggest that the kallikrein may be involved in prostate cancer development; however, there has been very little other progress toward its biochemical characterization or elucidation of its true physiological role.

8p system191 01-06-2013 45 2   Download

The present work reports isolation and characterization of a highly glycosylated protein from bovine milk fat globule membranes, known as PAS III. Partial amino-acid sequencing of the purified protein allowed construction of degenerate oligonucleotide primers, enabling isolation of a full-length cDNA encoding a protein of 330 amino-acid residues. N-terminal amino-acid sequencing of derived peptides and the purified protein confirmed 76% of the sequence and demonstrated presence of a cleavable signal peptide of 23 residues, leaving a mature protein of 307 amino acids.

9p system191 01-06-2013 50 4   Download

Penaeidins are a family of antimicrobial peptides constitutively produced and stored in the haemocytes of penaeid shrimp. In response to microbial stimulation, they are released into the blood circulation and they further attach to shrimp cuticle surfaces through a chitin-binding property. In the present paper, we have analysed their expression, regulation and distribution in shrimp tissues in response to experimental microbial challenge.

12p system191 01-06-2013 40 6   Download

Selenoprotein P (SeP) is a plasma protein that contains up to 10 selenocysteine residues and accounts for about 50% of total selenium in human plasma. We have previously shown that SeP expression in the human liver cell line HepG2 is inhibited by transforming growth factor (TGF)-b1 on a transcriptional level. Smad proteins are the transcriptional mediators of TGF-b signalling and putative Smad-binding elements (SBE) comprising the core sequence CAGACA are present at two positions in the SeP promoter....

6p system191 01-06-2013 35 4   Download

Departamento de Fisiologıa de la Nutricion, Instituto Nacional de Ciencias Medicas y Nutricion ‘Salvador Zubiran’, Mexico; ´ ´ ´ ´ ´ Departamento de Patologıa Experimental, Instituto Nacional de Ciencias Medicas y Nutricion ‘Salvador Zubiran’,Mexico; 3Department of Biochemistry, Wake Forest University Medical Center, Winston-Salem, North Carolina, USA Branched chain amino-acid aminotransferase (BCAT) activity is present in fetal liver but the developmental pattern of mitochondrial BCAT (BCATm) expression in rat liver has not been studied.

8p system191 01-06-2013 47 6   Download

The octadecaneuropeptide (ODN; QATVGDVNTDRPG LLDLK) and its C-terminal octapeptide (OP; RPGLLDLK), which exert anxiogenic activity, have been previously shown to increase intracellular calcium concentration ([Ca21]i) in cultured rat astrocytes through activation of a metabotropic receptor positively coupled to phospholipase C. It has also been found that the [D -Leu5]OP analog possesses a weak antagonistic activity. The aim of the present study was to synthesize and characterize cyclic analogs of OP and [D -Leu5]OP.

13p system191 01-06-2013 32 4   Download

The involvement of the lysine residue present at the active site of Ehrlich ascites carcinoma (EAC) cell glyceraldehyde-3-phosphate dehydrogenase (Gra3P DH) was investigated by using the lysine specific reagents trinitrobenzenesulfonic acid (TNBS) and pyridoxal phosphate (PP). Both TNBS and PP inactivated EAC cell Gra3P DH with pseudo-first-order kinetics with the rate dependent on modifier concentration. Kinetic analysis, including a Tsou plot, indicated that both TNBS and PP apparently react with one lysine residue per enzyme molecule.

8p system191 01-06-2013 37 3   Download

The isoprenoid methyl farnesoate (MF) has been implicated in the regulation of crustacean development and reproduction in conjunction with eyestalk molt inhibiting hormones and ecdysteroids. Farnesoic acid O-methyltransferase (FAMeT) catalyzes the methylation of farnesoic acid (FA) to produce MF in the terminal step of MF synthesis. We have previously cloned and characterized the shrimp FAMeT. In the present study, recombinant FAMeT (rFAMeT) was produced for bioassay and antiserum generation.

9p research12 01-06-2013 41 3   Download

Angiotensin converting enzyme (ACE) was already discovered in insects in 1994, but its physiological role is still enigmatic. We have addressed this problem by purifying four new ACE substrates from the ovaries of the grey fleshfly, Neobellieria bullata. Their primary structures were identified as NKLKPSQWISLSD (Neb-ODAIF-11)13), NKLKPSQWI (Neb-ODAIF-11)9), SLKPSNWLTPSE (Neb-ODAIF-2) and LEQIYHL. Database analysis showed significant homology with amino acid sequence stretches as present in the N-terminal part of several fly yolk proteins....

9p research12 01-06-2013 36 3   Download

We report the cloning and functional charaterization of the full-length cDNA and gene encoding a Toxoplasma gondii DNA repair enzyme designated TgDRE. The gene is composed of three exons separated by two introns of 780 and 630 bp, and encodes a protein with a predicted molecular mass of 49.6 kDa. The native TgDRE protein, with a molecular mass of 60 kDa, is only detected in the virulent tachyzoite stage of T. gondii. However, the transcript is present in both asexual parasite stages, virulent tachyzoite and avirulent encysted bradyzoite. ...

9p research12 01-06-2013 33 5   Download

The Saccharomyces cerevisiae type 2A protein phosphatase (PP2A) Pph22p differs from the catalytic subunits of PP2A (PP2Ac) present in mammals, plants and Schizosaccharomyces pombe by a unique N-terminal extension of approximately 70 amino acids. We have overexpressed S. cerevisiae Pph22p and its N-terminal deletion mutant DN-Pph22p in the GS115 strain of Pichia pastoris and purified these enzymes to apparent homogeneity.

11p research12 01-06-2013 45 5   Download