Laredoan Daniel Muñoa presents nano-technology abstract
At 103rd meeting of American Society for Microbiology in D.C.

By María Eugenia Guerra

Recent Texas A & M International University graduate Daniel Muñoa recently presented his abstract entitled "Modifying Viral Protein for Assembling Nano-Sized Constructs" at the May 23 General Meeting of the American Society for Microbiology (ASM) in Washington, D.C.
Muñoa wrote the abstract after research he conducted as a Minority Undergraduate Research Fellow at the Tufts Biomedical Research Center in the summer of 2002. He interned under the tutelage of Dr. Eddie Goldberg at Tufts from May 15 to August 08, 2002. Goldberg is a professor of Molecular and Microbiology at Tufts School of Medicine.
At the 103rd General Meeting Muñoa participated in a three-hour poster session encompassing the research he conducted at Tufts. About 20,000 scientists and doctors from around the world convened for the meeting at the new Washington Convention Center. "There were a lot of hardcore lab rats in attendance as well," Muñoa said, adding, "I saw it as an incredible gathering of the minds."
Presenting authors at the ASM's 103rd meeting were categorized into four Divisional Groups, each having numerous lettered divisions. Muñoa presented in the Molecular Microbiology, Physiology, and Virology Group, the Bacteriophage Division.
According to Muñoa, "Hundreds of millions of dollars are being spent yearly on innovative techniques to advance the rapidly growing field of nanotechnology, which is revolutionizing many industries by redesigning and building various existing functional systems such as robotics, computers, and even micro-scale laboratories from a much smaller order of magnitude (10-9-10-10 m) than what is currently attainable; e.g., computer transistors are currently assembled at the micron level, or 10-6 m."
Muñoa said that techniques in design of applicable systems in nanotechnology can be classified into two categories. Top-down techniques offer a traditional approach to design of a system; use machining and other etching techniques for design of micro-sized constructs. The major setback of top-down technique is difficulty to direct assembly beyond 10-6m (micron). Bottom-up techniques offer a design and construction of functional systems at the molecular level, well beyond the micron (micrometer). "Unfortunately," said Muñoa, "most existing applications of nanotechnology utilize top-down techniques for assembly. To date, the only existing manufacturing plant functioning at the order of the nanometer is the living cell, be it prokaryotic or eukaryotic. The manufacturing plants in both prokaryotic and eukaryotic cells are exploited during viral infection for the purpose of reproduction and subsequent host infection.
"All living organisms have a genetic foundation upon which they exist," he continued. "For humans, that genetic foundation is the 46 chromosomes that comprise the human genome. For simpler forms of life such as bacteria, the genetic information is so small that there are no visible chromosomes present. Instead, loosely double-stranded DNA aggregates in a certain region within the bacterial cell. In any event, the methods by which DNA is accessed are very similar across all spectrums of life. The central dogma in genetics is that DNA, in the form of functional genes, are 'transcribed' to mRNA via enzymatic reactions and subsequently 'translated' into polypeptides (proteins), comprised of amino acids. Proteins are the most diverse class of macromolecules and fall under two broad categories, functional or structural proteins. An example of a structural protein would be the hair that grows on mammals, while functional proteins are better known as enzymes. One example of an enzyme is DNA polymerase, and its function is to replicate DNA during mitotic events. Many of the experiments that I conducted were based on principles and protocols from the field of molecular biotechnology. This is a fast growing branch of molecular biology that has branched off into several sub-groups of molecular-based studies such as proteomics, genomics, and bioinformatics.
"To make a long, scientific story short," said Muñoa, "Dr. Goldberg's intentions are to utilize portions of existing biological structures (viruses) as a novel source of building material for assembling functional constructs at the order of the nanometer. In particular, segments of tail fibers from T4 bacteriophage are the focus of Dr. Goldberg's research. Tail fibers can be thought of as two-by-fours that you use to build a house, except these two-by-fours exist at a level many orders of magnitude smaller that what we are accustomed to thinking of. The size of tail fibers and viruses are measured between the orders of the angstrom and the nanometer, which are 10-10 m and 10-9 m, respectively. However, in order to have nanoscopic "two-by-fours" capable of directed self-assembly in vitro, there are several hurdles that need to be overcome. One of those hurdles was the focus of my summer fellowship in 2002 at Tufts," said Muñoa.
"The tail fiber is a structural protein that functions as an anchoring mechanism for the virus to attach itself to the surface of a bacterial cell prior to injection of viral DNA," said Muñoa. "Dr. Goldberg believes that tail fiber segments encoded by gene 37 are rigid and durable enough to be used as a source of building material based on the following premise: Tail fibers withstand considerable amounts of mechanical stress in bringing the viral head and tail close enough to the outer surface of a bacterial cell for the deployment of a needle-like shaft encased within the tail. This tail penetrates the bacterium's cellular membrane, allowing for the introduction of viral DNA stored within the viral head to be transmitted to the host. The cellular machinery of bacterial cells cease normal function and begin to 'transcribe' and 'translate' viral DNA, producing new viruses that will ultimately burst out of the host, causing its death, and probe the surrounding environment for other bacterial cells to invade."
Muñoa said that during his fellowship at Tufts, he genetically modified gene 37 of T4 phage. Gene 37 encodes the terminal segments of tail fibers in T4 phage. "The purpose of my experiments were to indirectly analyze the three-dimensional structure of tail fiber segment 37 by searching for functional phage with shortened tail fibers resulting from nested, unidirectional deletions I performed on gene 37. The deletion junction formed in tail fiber segment 37 as a result of deleted segments within gene 37 are currently being tested in several ways to determine if they serve as suitable sites for encoding binding residues that would permit assembly of nano-sized constructs solely based on tail fiber segments attached to one another and/or a functional moiety. However, I was not involved in this area of study and cannot elaborate any further on assembly of structures," said Muñoa.
"To date, only one T4 phage has been produced that is missing close to a thousand base pairs from gene 37 by Dr. Paul Hyman, leading scientific investigator at Nanoframes, a bio-tech company based out of Boston owned by Dr. Goldberg. This means that the resultant tail fiber segment is missing approximately 333 amino acid residues when translated from mRNA. The underlying concept in determining the extent of amino acid residues missing from the resultant, altered, protein structure is that mRNA is read three nucleotide bases at a time in frames known as codons. There are 64 possible codons. The majority of the 64 codons encode for, with varying redundancy, the 20 amino acids necessary for protein synthesis in living organisms. A few of the 64 possible codons serve as start or stop signals for enzymatic machinery involved in protein synthesis," said Muñoa.
Muñoa completed the remaining phase of his research at TAMIU in the fall 2002 and spring 2003 semesters. "I worked on my project up until April of 2003, but was unable to produce any phage exhibiting shortened tail fibers," he said. "I did screen close to 1,000 recombinant colonies of bacteria. Each colony possessed various sized deletions of gene 37 loaded on a plasmid that is subsequently uploaded into the phage's genome via a crossing over event. This means that segments of DNA homologous to one another arising from different templates are exchanged. In this case, altered gene 37 on the plasmid would be exchange for gene 37 of the virus' genome. This is a classic model of gene transfer studied in genetics that viruses utilize to exchange genetic information with one another. Although I failed in yielding positive results, I must say that the knowledge and experienced gained in scientific endeavor during the summer of 2002 is truly priceless. Dr. Goldberg told me from the get-go, 'The odds are against you hitting a positive, but then again what's important here is that you learn something from setting all your stuff up on the work bench and at the end of the day, placing yourself in position to generate a positive result.'"
While studying for the medical school entry exam, Muñoa continues conducting independent research with his mentor Dr. Mario García Ríos in the TAMIU biology department.

faculty member of the year

By Paul de la Peña-Franceschi

Dr. Joseph Crabtree was named Laredo Community College (LCC) faculty member of the year at its recent 18th annual LCC Employment Appreciation Day. Dr. Crabtree currently directs the mixed choir and opera workshop programs at LCC and is also involved in many other musical activities in the community. Recently tenured, Dr. Crabtree received service recognition for five years. He was recognized for his dedication to students and his creative teaching abilities. He was also the school's nominee for the Golden Apple award.
Of the recognition accorded him by his peers, Dr. Crabtree said, "I feel exceedingly honored and humbled by this achievement and am committed to offering the best job I can for the students,"
According to Dr. Crabtree, he would not have pursued music were it not for the urging of his sister to drop computer class and go into music. She recognized in him, he said, a talent that was often overlooked in his middle school years. He was told in 1991 that his voice was too small for the stage and had zero performance potential. He was told to look for another line of work. Ignoring this assessment, he continued in music, landing a chorus role in the musical Guys and Dolls in a high school production in Ohio. He has since studied piano, violin, flute, and voice and has performed in church choirs most of his life. Heeding his sister's advice, he has now made a name for himself for achievements on and off campus and in concert halls on two continents.
Dr. Crabtree is an acclaimed performer on both sides of the Atlantic in over 25 leading operatic roles and as a choral conductor. A native of Columbus, Ohio, Crabtree received his Bachelor of Music degree in vocal performance from Ohio State University. During his studies there, he worked with such noted composers and conductors as Maurice Casey, James Gallagher, Norman Luboff, Robert Shaw, Alice Parker, William Mathias, Jester Hairston, and John Rutter. His singing engagements in Ohio included performing with the Cantari Singers of Columbus and the Lancaster Chorale, where his chorale work, "A Benediction," was featured in their tenth anniversary concert. He was the baritone soloist in Columbus' first performance of Puccini's Messe di Gloria.
In 1992, Dr. Crabtree moved to Austin to pursue Masters and Doctorate degrees in Voice Performance at the University of Texas at Austin. He became recognized as a comic bass-baritone, and was featured in such roles as Bartolo in The Marriage of Figaro, 80-year-old Simone in Gianni Schicchi, and Rapunzel's Prince/Wolf in Into the Woods. He credits William Lewis, an accomplished lifetime musician for at the New York Metropolitan Opera, for much of his training at the University of Texas. Lewis, also Artistic Director at the Austrian American Mozart Academy, directed Crabtree's stage presence and gave him numerous opportunities to perform and act. Voice coach Darlene Wiley to whom Crabtree refers to as "the goddess of voice," helped him master his voice through constant direction in master classes and vocal coaching.
While at UT Dr. Crabtree attended the Austrian American Mozart Academy in Salzburg, Austria, where he studied and performed the works of Mozart in Mozart's homeland. He excelled in Mozart's comedic roles -- Papageno, Leporello, Nardo, and Bartolo -- and performed these with the Amadeus Ensemble throughout Austria and Germany. He was also the bass soloist in performances of Mozart's C-Minor Mass and Requiem.
Adept at changing styles, Dr. Crabtree performed in early music productions of L'Ormindo by Cavalli with period instrumentation, as well as the 20th-century works of Leonard Bernstein and Dominic Argento. He was a featured soloist with the New Music Ensemble in Austin, where he performed John Harbison's Words from Patterson, with the composer in attendance. His broad range of musical work encompasses oratorio, art song, American musical theater, Viennese operetta, and opera, and he has even been featured in radio and TV commercials.
In 1997, Dr. Crabtree joined the faculty of Laredo Community College and now considers Laredo home. He spends his summers in Salzburg, where he serves on the faculty of the Austrian American Mozart Academy, and conducts the church music throughout the Salzkammergut area, performing both of Mozart's Litanies in B-flat, the Coronation Mass, and the Solemn Vespers. This summer he will make his conducting debut directing Mozart's first opera, Bastien und Bastienne and Der Schauspieldirektor.
Since his arrival in Laredo, Dr. Crabtree has supported the arts through recitals at the Center for the Arts, as well in his performances of Tevye in Fiddler on the Roof, and the Cowardly Lion in The Wizard of Oz, both staged by Laredo Institute for Theatrical Education (L.I.T.E.) Productions. He has been a soloist with both the Laredo Philharmonic Chorale and the Laredo Philharmonic Orchestra. At LCC he has directed six full opera productions including The Magic Flute, The Empresario, The Fantastiks, Ol'Maid and the Thief, Little Shop of Horrors, and The Pirates of Penzance. The mixed choir has performed the Liebeslieder waltzes by Brahms and most recently performed an evening of Schubert singing his Mass in F and Hirtenchor (the Shepard's Chorus).
Dr. Crabtree completed his Doctor of Musical Arts (D.M.A). in the spring of 2001. The title of his dissertation was "An Investigation and Analysis of Ned Rorem's Fables-Five Very Short Operas." He was then immediately hired as the artistic director and conductor of the Laredo Philharmonic Chorale (LPC).
Under his direction the LPC has experienced tremendous growth and challenge during the past two years in which he has directed a series of Broadway music, show tunes, and American patriotic hymns and ballads. He has also conducted the Mozart Requiem, Handel's Messiah, Ralph Vaughn Williams Five Mystical Songs, and Fáure's Requiem. This May the LPC returns to New York City's Carnegie Hall to perform Mozart's Requiem under the direction of John Rutter. Additionally, in June the women's chorus has been invited to perform in Monterrey performing Gustav Holst's The Planets, along with the Monterrey Philharmonic Orchestra under the direction of Maestro Carrasco.
Dr. Crabtree spends many hours with his students preparing and designing sets for local productions. He makes himself available for tutorial sessions and additional rehearsal times. His dry, yet cunning sense of humor always seems to lighten conversation and ease tensions.
Dr. Crabtree is a member of the National Association of Teachers and the International Thespian Society. His beautiful companion, wife Dana, is also an accomplished professional vocalist and music instructor at LCC and adjunct professor at Texas A&M International University. Dr. Crabtree serves on the L.I.T.E. board and is the organist at the First Presbyterian Church.
For more information about the fine and performing arts at Laredo Community College, call Dr. Crabtree at 721-5330.