A Novel Technique-Bone Splitting and Bone Grafting in an Hourglass-shaped Bone Following Distraction Osteogenesis.

AIM: We describe a novel technique of bone splitting and bone grafting in managing hypoplastic or hourglass-shaped regenerated bone in distraction osteogenesis. BACKGROUND: Hourglass-shaped regenerated bone is a potential complication during distraction osteogenesis which is vulnerable to fracture when loaded. Our novel technique overcomes this by increasing the diameter of new bone formation via bone splitting and bone grafting. CASE DESCRIPTION: We report three cases with hypoplastic regenerated bone following distraction osteogenesis. It was treated with bone splitting and bone grafting. Although one case was complicated with an iatrogenic transverse fracture during the surgery, all three cases achieved the goal of increasing bone diameter during the subsequent consolidation phase. CONCLUSION: This relatively simple and novel surgical intervention can overcome the hourglass-shaped appearance, thus preventing potential fracture. CLINICAL SIGNIFICANCE: We emphasise the importance of identifying hypoplastic regenerate bone before the consolidation phase of distraction osteogenesis. The novel technique described is a simple surgical intervention which can prevent potential fracture through the newly formed bone. HOW TO CITE THIS ARTICLE: Lee HS, Tasarib R, Hamzah FC, et al. A Novel Technique-Bone Splitting and Bone Grafting in an Hourglass-shaped Bone Following Distraction Osteogenesis. Strategies Trauma Limb Reconstr 2020;15(3):175-178.

Computational study of bindings of olive leaf extract (OLE) to HIV-1 fusion protein gp41.

Recent experimental study found that OLE (olive leaf extract) has anti-HIV activity by blocking the HIV virus entry to host cells [Lee-Huang, S., Zhang, L., Huang, P.L., Chang, Y. and Huang, P.L. (2003) Anti-HIV activity of olive leaf extract (OLE) and modulation of host cell gene expression by HIV-1 infection and OLE treatment. Biochem. Biophys. Res. Commun. 307, 1029; Lee-Huang, S., Huang, P.L., Zhang, D., Lee, J.W., Bao, J., Sun, Y., Chang, Y.-Tae, Zhang, J.Z.H. and Huang, P.L. (2007) Discovery of small-molecule HIV-1 fusion and integrase inhibitors oleuropein and hydroxytyrosol. Biochem. Biophys. Res. Commun. 354, 872-878, 879-884]. As part of a joint experimental and theoretical effort, we report here computational study to help identify and characterize the binding complexes of several main compounds of OLE (olive leaf extract) to HIV-1 envelop protein gp41. A number of possible binding modes are found by docking oleuropein and its metabolites, aglycone, elenolic acid and hydroxytyrosol, onto the hydrophobic pocket on gp41. Detailed OLE-gp41 binding interactions and free energies of binding are obtained through molecular dynamics simulation and MM-PBSA calculation. Specific molecular interactions in our predicted OLE/gp41 complexes are identified and hydroxytyrosol is identified to be the main moiety for binding to gp41. This computational study complements the corresponding experimental investigation and helps establish a good starting point for further refinement of OLE-based gp41 inhibitors.

Anti-HIV agent MAP30 modulates the expression profile of viral and cellular genes for proliferation and apoptosis in AIDS-related lymphoma cells infected with Kaposi's sarcoma-associated virus.

The anti-HIV agent MAP30 (Momordica anti-HIV protein, 30 kDa) inhibits the proliferation of BC-2, an AIDS-related primary effusion lymphoma (PEL) cell line derived from an AIDS patient. BC-2 cells are latently infected with Kaposi's sarcoma-associated herpes virus (KSHV), also known as human herpes virus 8 (HHV8). We examined the effect of MAP30 on the expression of viral and cellular genes in BC-2 during latent and lytic states of the viral life cycle. By Northern analysis and RT-PCR, we found that MAP30 downregulates the expression of viral cyclin D (vCD), viral interleukin-6 (vIL-6), and viral FLIP (vFLIP), genes involved in cell cycle regulation, viral pathogenesis, and apoptosis. By pathway-specific cDNA microarray analysis, we found that BC-2 cells express high levels of egr-1, ATF-2, hsp27, hsp90, IkappaB, mdm2, skp1, and IL-2, cellular genes involved in mitogenesis, tumorigenesis, and inhibition of apoptosis in NFkappaB and p53 signaling pathways. These results define for the first time the specific cellular pathways involved in AIDS-related tumorigenesis and suggest specific novel targets for the treatment. Furthermore, we found that MAP30 downregulates the expression of egr-1, ATF-2, hsp27, hsp90, IkappaB, mdm2, and Skp1, while it upregulates the pro-apoptotic-related genes Bax, CRADD, and caspase-3. Thus, MAP30 modulates the expression of both viral and cellular genes involved in KS pathogenesis. These results provide valuable insight into the molecular mechanisms of MAP30 anti-KS action and suggest its utility as a therapeutic agent against AIDS-related tumors.

Inhibition of MDA-MB-231 human breast tumor xenografts and HER2 expression by anti-tumor agents GAP31 and MAP30.

GAP31 (Gelonium protein of 31 kDa) and MAP30 (Momordica protein of 30 kDa) are agents isolated from the medicinal plants Gelonium multiflorum and Momordica charantia, respectively. The current study was conducted to investigate the efficacy of GAP31 and MAP30 on estrogen-independent and highly metastatic human breast tumor MDA-MB-231 both in vitro and in vivo. The effect of these agents on the expression of breast tumor antigen HER2 (also known as neu or as c-erbB 2) was also examined. Treatment of MDA-MB-231 breast cancer cells with GAP31 and MAP30 resulted in inhibition of cancer cell proliferation as well as inhibition of the expression of HER2 gene in vitro. When MDA-MB-231 human breast cancer cells were transferred into SCID mice, the mice developed extensive metastases and all mice succumbed to tumor by day 46. Treatment of the human breast cancer bearing SCID mice with GAP31 or MAP30 at 10 micrograms/injection EOD for 10 injections resulted in significant increases in survival, with 20-25% of the mice remaining tumor free for 96 days. Thus, anti-tumor agents GAP31 and MAP30 are effective against human breast cancer MDA-MB-231 in vitro and in vivo. These agents may therefore be a potential therapeutic use against breast carcinomas.

Anti-HIV and anti-tumor protein MAP30, a 30 kDa single-strand type-I RIP, shares similar secondary structure and beta-sheet topology with the A chain of ricin, a type-II RIP.

MAP30 is a 30 kDa single-stranded, type-I ribosome inactivating protein (RIP) possessing anti-tumor and anti-HIV activities. It binds both ribosomal RNA and the HIV-1 long-terminal repeat DNA. To understand the structural basis for MAP30 activities, we undertook the study of MAP30 by solution NMR spectroscopy. We report nearly complete 1H, 13C, and 15N chemical shift assignments of its 263 amino acids. Based upon an analysis of secondary 13C chemical shifts, 3J(HNHA) coupling constants, hydrogen exchange data, and nuclear Overhauser effect patterns, we find that the secondary structure and beta-sheet topology of MAP30 are very similar to those of the ricin A chain, a subunit of the well-known type-II RIP, even though two proteins display distinct activities. We therefore suggest that MAP30 and ricin A chain share a similar three-dimensional fold, and that the reported functional differences between two proteins arise primarily from differences in local three-dimensional structure and other structural properties such as surface electrostatic potentials.

Solution structure of anti-HIV-1 and anti-tumor protein MAP30: structural insights into its multiple functions.

We present the solution structure of MAP30, a plant protein with anti-HIV and anti-tumor activities. Structural analysis and subsequent biochemical assays lead to several novel discoveries. First, MAP30 acts like a DNA glycosylase/apurinic (ap) lyase, an additional activity distinct from its known RNA N-glycosidase activity toward the 28S rRNA. Glycosylase/ap lyase activity explains MAP30's apparent inhibition of the HIV-1 integrase, MAP30's ability to irreversibly relax supercoiled DNA, and may be an alternative cytotoxic pathway that contributes to MAP30's anti-HIV/anti-tumor activities. Second, two distinct, but contiguous, subsites are responsible for MAP30's glycosylase/ap lyase activity. Third, Mn2+ and Zn2+ interact with negatively charged surfaces next to the catalytic sites, facilitating DNA substrate binding instead of directly participating in catalysis.

The antiviral agents, MAP30 and GAP31, are not toxic to human spermatozoa and may be useful in preventing the sexual transmission of human immunodeficiency virus type 1.

OBJECTIVE: To investigate the effects of two virucidal compounds, MAP30 (Momordica anti-human immunodeficiency virus [HIV] protein; molecular weight, 30 kd) and GAP31 (Gelonium anti-HIV protein; molecular weight, 31 kd), obtained from Momordica charantia and Gelonium multiflorum, respectively, on the motility and vitality of human spermatocytes. DESIGN: Prospective, controlled study. SETTING: New York University School of Medicine. PATIENT(S): Ten healthy men undergoing evaluation for infertility provided 10 semen specimens. INTERVENTION(S): Human sperm were treated with the anti-HIV agents, MAP30 and GAP3 1. Nonoxynol-9, a commonly used spermicide, and phosphate-buffered saline were used as the positive and negative controls, respectively. MAIN OUTCOME MEASURE(S): The motility and vitality of human spermatocytes treated with MAP30 and GAP31 at doses that inhibit HIV-1 and herpes simplex virus. RESULT(S): MAP30 and GAP31 did not inhibit the motility or vitality of human sperm cells over a dose range of 100-0.1 microg/mL, whereas nonoxynol-9 demonstrated spermicidal action on all 10 samples over the same dose range. CONCLUSION(S): The antiviral agents, MAP30 and GAP31, were not toxic to human sperm cells at the doses at which they inhibit HIV-1 and herpes simplex virus. They had no effect on the motility of spermatozoa, even at a dose of 1,000 times the maximum effective concentration. These results indicate that MAP30 and GAP31 may be useful as nonspermicidal protection against sexually transmitted diseases.

Proteolytic fragments of anti-HIV and anti-tumor proteins MAP30 and GAP31 are biologically active.

We analyzed the structural and functional organization of anti-HIV and anti-tumor proteins MAP30 and GAP31 by limited proteolysis with endopeptidases Lys-C and Glu-C (V8). MAP30 and GAP31 are resistant to proteolytic digestion under conditions of as much as 5% (w/w) proteases. In the presence of 10% (w/w) protease, the central regions of the proteins are still resistant to proteolysis, whereas the N- and C-termini are accessible. Peptide fragments were purified by FPLC on Superdex 75 columns, characterized by gel electrophoresis, identified by amino acid sequencing, and analyzed for anti-HIV, anti-tumor, and other biochemical activities. We report here that limited proteolysis yields biologically active fragments of both MAP30 and GAP31. These fragments are active against HIV-1 and tumor cells with EC(50)s in the sub-nanomolar ranges, 0.2-0.4 nM. At the dose levels used in the assays, little cytotoxicity to normal cells was observed. In addition, these fragments remain fully active in HIV-integrase inhibition and HIV-LTR topological inactivation, but not ribosome inactivation. These results demonstrate that the antiviral and anti-tumor activities of MAP30 and GAP31 are independent of ribosome inactivation activity. In addition, we demonstrate that portions of the N- and C-termini are not essential for antiviral and anti-tumor activities, but do appear to be required for ribosome inactivation. These results may provide novel strategies for rational design and targeted development of mimetic antiviral and anti-tumor therapeutics.

Measurement of 3hJNC' connectivities across hydrogen bonds in a 30 kDa protein.

A method is described which permits detection of 3hJNC' scalar couplings across hydrogen bonds in larger, perdeuterated proteins. The experiment is demonstrated for the uniformly 2H/13C/15N-enriched 30 kDa ribosome inactivating protein MAP30. The 3hJNC' interactions are smaller than 1 Hz, but their detection in an HNCO experiment is made possible through the use of constructive interference between the 15N chemical shift anisotropy and 1H-15N dipole-dipole relaxation mechanisms in a manner similar to that of recently proposed TROSY schemes. Sensitivity of the HNCO experiment depends strongly on the 15N transverse relaxation rate of the downfield 15N multiplet component and on the amide proton T1. In perdeuterated MAP30 at 40 degrees C, the average TROSY T2 was 169 ms at 750 MHz 1H frequency, and a wide range of longitudinal relaxation rates was observed for the amide protons.

Lysozyme and RNases as anti-HIV components in beta-core preparations of human chorionic gonadotropin.

Human chorionic gonadotropin (hCG) preparations contain activity against HIV type 1 (HIV-1). However, there has been controversy about whether some biological activities of hCG beta-subunit (hCGbeta) preparations are caused by the beta-subunit itself or other proteins present in the preparations. We report here the purification, characterization, and identification of three enzymes with anti-HIV activity present in the beta-core fraction of hCGbeta prepared from the urine of pregnant women. The N-terminal amino acid sequence of one protein is identical to human urinary lysozyme C, and those of the other two are identical to human RNase A and urinary RNase U. We thus refer to these proteins as AVL (antiviral lysozyme) and AVR (antiviral RNases). In addition to HIV-1 inhibition, AVL is capable of lysing Micrococcus lysodeikticus. AVR digests a variety of RNA substrates, including RNA from HIV-1-infected cells. We also find that lysozyme from chicken egg white, human milk, and human neutrophils and RNase A from bovine pancreas possess activity against HIV-1. These findings may offer additional strategies for the treatment of HIV-1 infection.

The activity of plant-derived antiretroviral proteins MAP30 and GAP31 against herpes simplex virus in vitro.

We examined the effect on anti-HIV proteins MAP30 and GAP31, from Momordica charantia and Gelonium multiflorum, on the infection and replication of Herpes Simplex Viruses (HSV). Human lung WI-38 fibroblasts cultured in the presence of tenfold dilutions of MAP30 or GAP31 were exposed to HSV and viral yield was measured at 24-48 hours by ELISA. The effective concentrations for 50% inhibitions (EC50) were 0.1-0.2 microM for HSV-2, and 0.3-0.5 microM for HSV-1 for MAP30 and GAP31, respectively. In comparison, the EC(50) for acyclovir (ACV), a commonly used anti-HSV drug, was 0.2 and 1.7 microM for HSV-2 and HSV-1, respectively. The cytotoxicity of all three antivirals was negligible and comparable. However, the antiherpetic activity of the plant proteins against acyclovir-resistant strains was two to three logs more potent than ACV. These results suggest that MAP30 and GAP31, previously shown to be active against HIV, may be useful for the therapy of herpesvirus infections.

Inhibition of the integrase of human immunodeficiency virus (HIV) type 1 by anti-HIV plant proteins MAP30 and GAP31.

MAP30 (Momordica anti-HIV protein of 30 kDa) and GAP31 (Gelonium anti-HIV protein of 31 kDa) are anti-HIV plant proteins that we have identified, purified, and cloned from the medicinal plants Momordica charantia and Gelonium multiflorum. These antiviral agents are capable of inhibiting infection of HIV type 1 (HIV-1) in T lymphocytes and monocytes as well as replication of the virus in already-infected cells. They are not toxic to normal uninfected cells because they are unable to enter healthy cells. MAP30 and GAP31 also possess an N-glycosidase activity on 28S ribosomal RNA and a topological activity on plasmid and viral DNAs including HIV-1 long terminal repeats (LTRs). LTRs are essential sites for integration of viral DNA into the host genome by viral integrase. We therefore investigated the effect of MAP30 and GAP31 on HIV-1 integrase. We report that both of these antiviral agents exhibit dose-dependent inhibition of HIV-1 integrase. Inhibition was observed in all of the three specific reactions catalyzed by the integrase, namely, 3' processing (specific cleavage of the dinucleotide GT from the viral substrate), strand transfer (integration), and "disintegration" (the reversal of strand transfer). Inhibition was studied by using oligonucleotide substrates with sequences corresponding to the U3 and U5 regions of HIV LTR. In the presence of 20 ng of viral substrate, 50 ng of target substrate, and 4 microM integrase, total inhibition was achieved at equimolar concentrations of the integrase and the antiviral proteins, with EC50 values of about 1 microM. Integration of viral DNA into the host chromosome is a vital step in the replicative cycle of retroviruses, including the AIDS virus. The inhibition of HIV-1 integrase by MAP30 and GAP31 suggests that impediment of viral DNA integration may play a key role in the anti-HIV activity of these plant proteins.

Anti-HIV and anti-tumor activities of recombinant MAP30 from bitter melon.

MAP30 is an anti-HIV plant protein that we have identified and purified to homogeneity from bitter melon (Momordica charantia). It is capable of acting against multiple stages of the viral life cycle, on acute infection as well as replication in chronically infected cells. In addition to antiviral action, MAP30 also possesses anti-tumor activity, topological inactivation of viral DNA, inhibition of viral integrase and cell-free ribosome-inactivation activities. We have cloned and expressed the MAP30 gene. The objective of this study is to characterize recombinant MAP30 (re-MAP30) and to determine its anti-HIV, anti-tumor and other activities. We report here that re-MAP30 inhibits HIV-1 and certain human tumors to the same extent as its native counterpart, natural MAP30 (nMAP30). The anti-HIV activity was measured by quantitative focal syncytium formation on CEM-ss cell monolayers, viral core protein p24 expression and viral-associated reverse transcriptase activity in HIV-1-infected H9 cells. The anti-tumor activity was measured by metabolic labeling of protein synthesis in tumor cells. In the dose range of the assay, re-MAP30 exhibits little toxicity to the uninfected viral target cells and other normal human cells. Identical to nMAP30, re-MAP30 is also active in topological inactivation of viral DNA, inhibition of viral DNA integration and cell-free ribosome inactivation. The cloning and expression of the gene encoding biologically active re-MAP30 provides an abundant source of homogeneous material for clinical investigations, as well as structure-function studies of this novel antiviral and anti-tumor agent.

Protective effect of interferon-alpha against cell-mediated human immunodeficiency virus transmission resulting from coculture of infected lymphocytes with fetal trophoblasts.

The hypothesis that the low transmission rate of HIV in utero may be due, in part, to the protective effect of IFN-producing placental trophoblasts was explored in vitro. The model consisted of H9 lymphocytes, as surrogates of maternal HIV-infected T cells, incubated for 3 h with JEG-3 trophoblasts in the presence of 10-fold dilutions of leukocyte-derived IFN-alpha (from 1000 to 0.1 IU/ml). The dose effect was monitored either directly, by measuring the levels of proviral DNA by PCR after a single round of infection, or indirectly, by coculturing infected JEG-3 with cord blood-derived MT-4 lymphocytes and determining the levels of p24 production by ELISA. Both assays revealed a dose-dependent blocking effect of IFN-alpha on cell-mediated HIV transmission. The complete inhibition of HIV infection was observed in the presence of 100 IU IFN-alpha. The efficacy of such a low dose could not be attributed to insufficient viral load because up to 10(8) infectious particles could be transmitted during cell-cell contact. An adhesion assay ruled out the possibility that IFN-alpha acts through prevention of lymphocyte-trophoblast contact. The results suggest that physiologic levels of IFN-alpha, present in the placental environment, may contribute to the protection of the fetus against HIV infection.

Acrosin inhibitor, 4'-acetamidophenyl 4-guanidinobenzoate, an experimental vaginal contraceptive with anti-HIV activity.

Serine proteases are involved in a wide variety of seemingly unrelated physiological functions including capacitation of the spermatozoa and potentiation of human immunodeficiency virus (HIV) infection. The experimental vaginal contraceptives derived from 4-guanidinobenzoic acid act through inhibition of acrosin--a serine protease from the sperm. The serial ten-fold dilutions of 4'-acetamidophenyl 4-guanidinobenzoate (AGB) were tested in vitro for the effect against HIV infection by assaying the suppression of de novo p24 synthesis in virus-inoculated MT-4 T lymphocytes. The results reveal that complete inhibition of HIV occurred at 100 micrograms/ml--a dose corresponding to previously reported concentrations responsible for preventing fertilization in rabbits. These findings suggest that serine protease inhibitors and in particular the guanidinobenzoates, reported to be up to 100-fold more potent and less irritating than nonoxynol-9, can be potentially operative against sexual transmission of HIV.

PIP: Biochemists at the New York University Medical Center assayed suppression of de novo p24 synthesis in HIV-inoculated MT-4 T lymphocytes to compare the effects of 4'-acetamidophenyl 4-guanidinobenzoate hydrochloride (AGB), Nalpha-p-tosyl-L-lysine chloro methyl ketone (TLCK), and nonoxynol-9 against HIV infection. Specifically, they wanted to determine the possibility that AGB, an antagonist of acrosin (a serine protease critical to sperm capacitation and to fortification of HIV infection), could launch a new class of vaginal spermicidal agents with anti-HIV activity. 100 mcg/ml AGB (a nontoxic dose) induced complete inhibition of HIV but did not affect the viability of MT-4 T lymphocytes. AGB had a 100 times more potent effect against HIV and had a lower cytotoxic effect (i.e., less irritating) than nonoxynol 9. These findings suggest that serine protease inhibitors, especially the guanidinobenzoates, have the potential to be effective vaginal contraceptives with anti-HIV activity.

Potentiation of anti-HIV activity of anti-inflammatory drugs, dexamethasone and indomethacin, by MAP30, the antiviral agent from bitter melon.

MAP30 is an antiviral protein from bitter melon (Momordica charantia). The enhancement of weak HIV antagonists, dexamethasone and indomethacin, by MAP30 has been examined by measuring the reduction in p24 expression in acutely infected MT-4 lymphocytes. In the presence of 1.5 nM MAP30 the IC50 dose of dexamethasone and indomethacin has been lowered, without concurrent cytotoxicity, at least a thousand-fold to 10(-7) M and 10(-8) M, respectively. This observation indicates that MAP30, a multifunctional antiviral plant protein capable of topological inactivation of viral DNA and specific cleavage of 28 S ribosomal RNA, may regulate HIV replication in concert with steroid and non-steroidal inhibitors of prostaglandin synthesis. The results suggest that use of MAP30 in combination with low pharmacological doses of dexamethasone and indomethacin may improve the efficacy of anti-HIV therapy.

The non-steroidal anti-inflammatory drug, indomethacin, as an inhibitor of HIV replication.

Indomethacin, a common non-steroidal anti-inflammatory drug (NSAID), has been used to treat rheumatoid arthritis. Although indomethacin has also been used as an immunopotentiator and symptomatic NSAID in AIDS, its effect on HIV replication is unknown. MT-4 lymphocytes were inoculated with HIV in the presence of indomethacin and tested for p24 expression by ELISA. The 50% inhibition (IC50) was 10 microM, corresponding to plasma levels after administration of 50 mg oral indomethacin. The antiviral effect appears to be specific since no toxicity has been observed at the IC50 dose, and unrelated NSAIDs have not shown the activity at clinical doses. Indomethacin may, thus, represent a new class of anti-HIV drug.

Anti-HIV effect of beta subunit of human chorionic gonadotropin (beta hCG) in vitro.

Human chorionic gonadotropin (hCG)--a pregnancy-associated immunomodulating hormone--has been recently shown in vitro to suppress reverse transcriptase activity in chronically HIV-infected lymphocytes and monocytes and to block viral transmission resulting from cell-cell contact between virus-carrying lymphocytes and placental trophoblasts. In further pursuit of the query into the mechanism of action, purified alpha and beta subunits of hCG were tested for the inhibition of p24 gag protein synthesis in virus-producing ACH-2 lymphocytes and U1 monocytes. Unlike the alpha subunit, beta-hCG displayed a distinct U-shaped dose response, characteristic of the effect of dimer hCG. Maximum inhibition of viral expression has been achieved at 10-100 ng/ml, the concentration corresponding to blood levels of beta-hCG in pregnant women. The doses that were several logs higher of normal levels seemed to increase viral production in monocytes. The data presented supports our original observations regarding the effect of intact hCG on HIV replication. While the mechanism of action remains to be established, the results suggest that the virus-interfering activity of hCG is determined by hormone-specific beta chain but not by the alpha subunit--shared with the family of glycoprotein hormones from the pituitary--follicle-stimulating hormone, luteinizing hormone and thyrotropin.

Human immunodeficiency virus type 1 (HIV-1) inhibition, DNA-binding, RNA-binding, and ribosome inactivation activities in the N-terminal segments of the plant anti-HIV protein GAP31.

GAP31 (gelonium anti-HIV protein of 31 kDa) is an anti-HIV protein which we have identified and purified from a medicinal plant, Gelonium multiflorum. It is capable of inhibiting HIV-1 infection and replication. GAP31 also exhibits DNA topoisomerase inhibitor activity and RNA N-glycosidase activity. The ability of GAP31 to interrupt both DNA and RNA functions may be related to its multiple antiviral actions. To define the roles of these activities in the anti-HIV action of GAP31, a series of peptides corresponding to the N-terminal segment of GAP31 were synthesized and assayed for the aforementioned activities of the parent molecule. A 33-aa segment (KGATYITYVNFLNELRVKTKPEGNSHGIPSLRK) designated as K10-K42 is the shortest peptide necessary and sufficient for HIV-1 inhibition, DNA and RNA binding, and ribosome inactivation. The peptides were 2-5 orders of magnitude less active than GAP31. Truncation of 19 aa from the C terminus of K10-K42 resulted in the loss of all of these activities. On the other hand, deletion of N-terminal residues to give E23-K42 did not alter ribosome-inactivation activity but eliminated the other activities. These findings permit identification of a 7-aa sequence, KGATYIT, at the N terminus of K10-K42 that is critical for DNA binding and RNA binding, whereas a 9-aa sequence, SHGIPSLRK, at the C terminus is important to ribosome inactivation. Both regions contribute to anti-HIV activity. Histidine at position 35 is critical for all of these activities. The disparity of sequence requirements for inhibition of HIV infection and replication and for ribosome-inactivation activity suggests that the anti-HIV activity of most ribosome-inactivating proteins may not be the result of N-glycosidase activity alone. Mapping the minimal domain of GAP31 offers insights into the rational design of molecular mimetics of anti-HIV drugs.