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Protein & Peptide Letters, Volume 9, No. 4, 2002

 

Contents

 

CRAMP Analog Having Potent Antibiotic Activity Without Hemolytic Activity Pp.275-282

Shin-Won Kang, Dong Gun Lee, Sung-Tae Yang, Yangmee Kim,  Jae Il Kim, Kyung-Soo Hahm & Song Yub Shin

[Abstract]

 

Isolation and Characterization of an Angiogenin-Like Protein from Goat Plasma Pp.283-288

Tushar Kanti Maiti and Swagata Dasgupta

[Abstract]

 

Cleavage Specificities of Aspartic Proteinases toward Oxidized Insulin B Chain at Different pH Values Pp.289-294

Senarath B. P. Athauda,  and Kenji Takahashia

[Abstract]

 

Functional Expression of Human Methionine Aminopeptidase Type 1 in Saccharomyces cerevisiae Pp.295-303

Benjamin Dummitt, Ying Fei and Yie-Hwa Chang

[Abstract]

 

Misconceptions Arising from a Sign Discrepancy in Thermodynamic Data for the Gibbs  Free Energy Profile of Ribonuclease A Pp.305-313

Paul W. Chun

[Abstract]

 

A Prionogenic Peptide Derived from Sup35 Can Force the Whole GST Fusion Protein to Show Amyloid Characteristics Pp.315-321

Young Kee Chae, Kyoung Suk Cho, Woochun Chun

[Abstract]

 

Production of Recombinant Human Calcitonin from Silkworm (B. mori) Larvae Infected by Baculovirus Pp.323-329

Guan-zhen Yang, Zhen-zhen Chen, Da-fu Cui, Bo-liang Li, Xiang-fu Wu

[Abstract]

 

Isolation and Partial Characterisation of a Protein from Buck Seminal Plasma (Capra hircus),Homologous to Spermadhesins Pp.331-335

D.I.A. Teixeira, B.S. Cavada, A.H. Sampaio, A. Havt, C. Bloch Jr., M.V.Prates,F.B.M.B. Moreno, E.A.Santos, C.A.A.Gadelha, T.S.Gadelha, F.S.M.Crisóstomo, V. J. F.  Freitas,

[Abstract]

 

Biophysical Evidence of Lipid and Carbohydrate Binding Activities of Shrimp High Density Lipoprotein/? Glucan Binding Protein Pp.337-344

Claudia L. Vargas-Requena, Alejandra Hernández-Santoyo, Gloria Yepiz-Plascencia, Francisco Vargas-Albores, Inocencio Higuera-Ciapara, Adela Rodríguez-Romero, José L. Soulages and Rogerio R. Sotelo-Mundo,

[Abstract]

 

A Convenient Method for the Synthesis of C-Protected Esters of Boc-/Z-a,a- Dialkylamino Acidsby the Mixed Carboxylic – Carbonic Anhydride Method Pp.345-348

Vommina V. Suresh Babu, Kuppanna Ananda & Ganga–Ramu Vasanthakumar

[Abstract]

 

Effect of Phosphorylation and Aggregation on Tau Binding to DNA Pp.349-357

Qian Hua, and Rong-Qiao He

[Abstract]

 

Heavy Metal-Mediated Crystallization of Escherichia coli Phytase and Analysis of Bridging Interactions Pp.359-365

Daniel Lim and Zongchao Jia

[Abstract]

 

Abstracts

 

[Back to top] CRAMP Analog Having Potent Antibiotic Activity Without Hemolytic Activity

Shin-Won Kang, Dong Gun Lee, Sung-Tae Yang, Yangmee Kim,  Jae Il Kim, Kyung-Soo Hahm & Song Yub Shin

 

CRAMP-18 is an 18-residue functional region, corresponding to residues 16-33 of a mouse-derived antibiotic peptide CRAMP. To develop novel antibiotic peptides possessing strong antibiotic activity against bacterial, fungal and tumor cells without hemolytic activity, three analogs of CRAMP-18 were synthesized containing either Leu- or Lys-substitution. Leu-substitution ([L1, 8]-CRAMP-18) in the hydrophobic helix face of CRAMP-18 induced a dramatic increase in antibiotic activity without a significant increase in hemolytic activity. Lys-substitution ([K2, 13]-CRAMP-18 or [K9, 16]-CRAMP-18) in the hydrophilic helix face produced a smaller response. Therefore, [L1, 8]-CRAMP-18 may be an attractive candidate for developing novel peptide antibiotics.

 

[Back to top] Isolation and Characterization of an Angiogenin-Like Protein from Goat Plasma

Tushar Kanti Maiti and Swagata Dasgupta

 

Angiogenin, a blood vessel inducing protein has been implicated in wound healing and tumour progression. First isolated from human carcinoma cells, it has been subsequently isolated from human, bovine, rabbit, pig and mouse sera and bovine milk. This study reports the isolation of an angiogenic-like protein from goat plasma. The ribonucleolytic activity has been followed by yeast transfer RNA (tRNA) degradation using spectrophotometric and denaturing polyacrylamide gel electrophoresis methods. The chorioallantoic membrane (CAM) assay has been implemented to study its angiogenic activity. The presence of this protein has also been confirmed by strong binding with placental Ribonuclease Inhibitor (PRI).

 

[Back to top] Cleavage Specificities of Aspartic Proteinases toward Oxidized Insulin B Chain at Different pH Values

Senarath B. P. Athauda,  and Kenji Takahashia

 

The cleavage specificities of typical aspartic proteinases: pepsin A, gastricsin, cathepsin D and rhizopuspepsin, were examined at different pH values with oxidized insulin B chain as a substrate with special attention to the specificities near neutral pH. Significant differences in relative specificity for scissile bonds were observed between pH 2.0 and 5.5-6.5, which may be partly related with the changes in dissociation states of the His and Glu residues in the substrate and the ionizable residues in the active site of each enzyme.

 

[Back to top] Functional Expression of Human Methionine Aminopeptidase Type 1 in Saccharomyces cerevisiae

Benjamin Dummitt, Ying Fei and Yie-Hwa Chang

 

We expressed recombinant human methionine aminopeptidase type 1 (MAP or MetAP) in a map1 null yeast strain to determine the extent of functional complementation between the two proteins. The human MetAP1 protein fully rescued the slow growth phenotype associated with deletion of yeast MetAP1, suggesting that the yeast and human MetAP1 proteins may have similar roles in vivo. Expression of human MetAP1 in yeast has significance in understanding the function of the human protein, studying its in vivo substrate specificity, and developing specific anti-fungal drugs to target yeast MetAP1.

 

[Back to top]Misconceptions Arising from a Sign Discrepancy in Thermodynamic Data for the Gibbs  Free Energy Profile of Ribonuclease A

Paul W. Chun

 

An apparent discrepancy in the data for the Gibbs free energy change as a function of temperature at different pHs, originally published by Brandts in 1965 and repeated by Brandts and Hunt in 1967 with an unexplained change in sign, has lead to close to 40 years of misguided thinking in examining the thermodynamics of protein unfolding, including the frequently promulgated idea of cold denaturation. We have carried out a detailed analysis based on the Planck-Benzinger approach, which is very powerful in clarifying the fundamental aspects of biochemical energetics.

 

[Back to top] A Prionogenic Peptide Derived from Sup35 Can Force the Whole GST Fusion Protein to Show Amyloid Characteristics

Young Kee Chae, Kyoung Suk Cho, Woochun Chun

 

A prion determining 7-mer peptide derived from Sup35 was fused to glutathione S transferase (GST). The fusion protein was successfully overexpressed in Escherichia coli, and purified by employing affinity chromatography.  Upon incubation, it showed substantial aggregation suggesting the formation of amyloid-like fibrils. Congo Red binding strongly suggested that the fusion protein formed amyloid-like fibrils. By considering the steric hindrance of GST, the b-sheet formation should be in the anti-parallel fashion.

 

[Back to top] Production of Recombinant Human Calcitonin from Silkworm (B. mori) Larvae Infected by Baculovirus

Guan-zhen Yang, Zhen-zhen Chen, Da-fu Cui, Bo-liang Li, Xiang-fu Wu

 

A synthetic modified gene encoding the human calcitonin analog (hmCT) was expressed by use of the baculovirus expression system. After injection with recombinant baculoviruses, the hmCT-GST fusion protein was produced within the silkworm larvae. The fusion protein was purified by affinity chromatography. Biological activity of hmCT for hypercalcemic effect was determined in normal rats.

 

[Back to top] Isolation and Partial Characterisation of a Protein from Buck Seminal Plasma (Capra hircus),Homologous to Spermadhesins

D.I.A. Teixeira, B.S. Cavada, A.H. Sampaio, A. Havt, C. Bloch Jr., M.V.Prates, F.B.M.B. Moreno, E.A.Santos, C.A.A.Gadelha,T.S.Gadelha, F.S.M.Crisóstomo, V. J. F.  Freitas

 

Spermadhesins are a family of secretory proteins expressed in the male genital tract of pig, horse and bull. Their function and structure have been widely studied, especially those isolated from boar. However, there are no data concerning spermadhesins isolated from buck. Buck seminal plasma was collected and subjected to ion exchange chromatography on DEAE-Sephacel column followed by chromatography in a C18 column coupled to a HPLC system. The purification of the protein was determined by SDS-PAGE and MALDI-TOF analysis exhibiting a molecular mass of 12.5 KDa and showed to be structurally homologous to spermadhesins from boar and stallion.

 

[Back to top] Biophysical Evidence of Lipid and Carbohydrate Binding Activities of Shrimp High Density Lipoprotein/β Glucan Binding Protein

Claudia L. Vargas-Requena, Alejandra Hernández-Santoyo, Gloria Yepiz-Plascencia, Francisco Vargas-Albores, Inocencio Higuera-Ciapara, Adela Rodríguez-Romero, José L. Soulages and Rogerio R. Sotelo-Mundo

 

Crustacean High Density Lipoprotein/b-Glucan Binding Protein (HDL/BGBP) has been studied due to its role in nutrition and immune response via activation of the defense cells (hemocytes) upon binding 1,3-D-b-glucan carbohydrates. In this study, HDL/BGBP was found to be composed mainly of beta sheets, as determined by circular dichroism. Lipoprotein aggregation resulted when HDL/BGBP interacted with phospolipid vesicles, laminaribiose (1,3-b-glucan disaccharide) or heparin. HDL/BGBP has similar dissociation constants for laminaribiose (Kd=22 mM) or heparin (Kd=46 mM) as determined by 90° light scattering.

 

[Back to top] A Convenient Method for the Synthesis of C-Protected Esters of Boc-/Z-a,a- Dialkylamino Acidsby the Mixed Carboxylic – Carbonic  Anhydride Method

Vommina V. Suresh Babu, Kuppanna Ananda & Ganga–Ramu Vasanthakumar

 

The synthesis of C-protected esters of Boc-/Z-a,a-dialkylamino acids is accomplished by using alkyl/aryl chloroformate in presence of DMAP as a catalyst. The reaction proceeds through mixed carboxylic–carbonic anhydride, which was monitored by IR. The reaction was clean and complete in about 2 hr. All the esters prepared have been obtained in good yield and are fully characterized.

 

[Back to top] Effect of Phosphorylation and Aggregation on Tau Binding to DNA

Qian Hua, and Rong-Qiao He

 

The potential function of neuronal tau was found by our recent studies on the effect of tau on the melting temperature of both calf thymus DNA and plasmid pBluescript-II SK (Hua and He, Chin. Sci. Bull. 2000, 45:999-1001). Herein we examined whether or not the interaction of tau with DNA was related to phosphorylation and aggregation. Tau, phosphorylated by neuronal cdc2-like kinase, associated with DNA as shown by electrophoretic mobility shift assay. Similar to native tau, phosphorylated tau could increase the melting temperature of calf thymus DNA. When tau was aggregated or treated with formaldehyde, neither native tau nor phosphorylated tau kept its ability to interact with DNA, suggesting that binding of tau to DNA was in an aggregation-dependent, and a phosphorylation-independent, manner.

 

[Back to top] Heavy Metal-Mediated Crystallization of Escherichia coli Phytase and Analysis of Bridging Interactions

Daniel Lim and Zongchao Jia

 

Escherichia coli phytase is a phosphatase that catalyzes the hydrolysis of phytic acid into inorganic phosphate and myo-inositol.  Two crystal forms of this enzyme were obtained in the presence of heavy metals.  Crystal forms I and II were obtained with the heavy atoms CdCl2 and HgCl2 and diffracted to 1.5 Å and 2.25 Å resolution, respectively. Hg2+ and Cd2+ both acted as molecular bridge(s), linking and stabilizing E. coli phytase in the unit cell, and played a crucial role in the crystallization of phytase by bridging neighbouring symmetry related molecules.