St. Davids, PA: Fire blight is one of the most significant threats to commercial apple and pear production worldwide, with economic losses exceeding $100 million annually in the United States alone. Fire blight is a major problem in apple orchards throughout Pennsylvania, which is the fourth largest producer of apples in the United States. This devastating disease is caused by the bacterial pathogen Erwinia amylovora, which is classified in the same family as a number of dangerous human pathogens. Genetic studies of fire blight have shown that one of the virulence factor proteins required for systemic spread of the pathogen within the vascular tissue of the tree is an enzyme known as HsvA*. Fire blight bacteria lacking HsvA are much less virulent and are unable to spread effectively within the tree during infection, suggesting that HsvA is an important weapon in the arsenal of the pathogen.
In order to understand how HsvA works at the molecular level, student researchers at Eastern University from the College of Arts and Sciences, the Templeton Honors College, and nearby Conestoga High School, working with Dr. Jeff Lawton in the Department of Chemistry, characterized the biochemical activity of HsvA and found that HsvA is an amidinotransferase enzyme that displays novel substrate specificity. Amidinotransferases are enzymes that move an amidino group – a molecular fragment consisting of a carbon atom connected to two NH2 groups – from a donor molecule (usually arginine) to an acceptor molecule. Dr. Lawton’s research group found that HsvA uses a class of small organic molecules known as polyamines as acceptor substrates, with putrescine and spermidine being the most highly preferred. In plants, polyamines are known to play an essential role in the mobilization of defenses during infection. The discovery that HsvA uses polyamines as a substrate suggests that fire blight may be working to disrupt polyamine-dependent signaling pathways in apple trees that are required for defense mechanisms to operate effectively, thereby securing an advantage for the pathogen during infection.
The results of this research were published in the Journal of Biological Chemistry, a leading journal in the biochemical field, appearing online on 9 November 2017 and in print on 29 December 2017**. This paper also presents the molecular structure of HsvA, determined using X-ray crystallography in collaboration with Dr. Sreejesh Shanker at Baylor College of Medicine in Houston, Texas. The details of the molecular structure provide important insights into the way in which HsvA is likely to interact with substrates during the amidinotransfer reaction. As Dr. Lawton observed, “the information that we have gained from this work provides a starting point for the rational design of inhibitors that could potentially serve as novel strategies for controlling fire blight in apple orchards by blocking the activity of this essential enzyme during infection. We have begun moving in this direction already.” The atomic coordinates of the HsvA structure are available in the Protein Data Bank under accession number 5WPI.
This project involved seven student collaborators working with Dr. Lawton at Eastern University. Vicki Wallace-Kneale ’15 was a biochemistry major in the College of Arts and Sciences whose work on the project was supported by funding from the HBE Foundation. Grace Schaefer ’16, Benjamin Barnhart ’15, Thomas Coyle ’15, and David Metzler ’08 were biochemistry majors and Templeton Honors College scholars. Dorsin Chang and Jeffrey Huang were students at Conestoga High School whose involvement in the project was supported by a STEM education grant that the Department of Chemistry received from the VWR Charitable Foundation. This work was also supported by a Cottrell College Science Award that Dr. Lawton received from Research Corporation for Science Advancement. Inquiries about this work may be directed to Dr. Lawton by email at jlawton@eastern.edu or by phone at 610-341-1545.
* Oh CS, Kim JF, and Beer SV. (2205). "The Hrp pathogenicity island of Erwinia amlovora and identification of three novel genes required for systemic infection." Molecular Plant Pathology 6(2), 125-138
** Shanker S, Schaefer GK, Barnhart BK, Wallace-Kneale VL, Chang D, Coyle TJ, Metzler, DA, Huang J, and Lawton JA (2017). "The virulence-associated protein HsvA from the fire blight pathogen Erwinia amylovora is a polyamine amidinotransferase." Journal of Biological Chemistry 292(52), 21366-21380.
