JAN 09, 1939 - JUL 13, 2020
After a prolonged illness, on July 13, 2020 our dear friend and colleague Dr. Jarmila Janatova died at her home in Salt Lake City, UT. With her passing, the complement field has lost an extraordinary biochemist, one who played a very prominent role in the discovery of the thioester bond in C3 and C4, the entity at the heart of their ability to covalently attach to foreign targets. Shortly before her death, the International Complement Society recognized Jarmila’s vital contribution to this important milestone in our field through her selection for one of the three inaugural Pioneering Women in Complement Research awards.
Although hailing from the relatively small town of Pisek, Czechoslovakia, Jarmila received all of her formal education in the culturally rich capital city, Prague. She received an M.Sc. in Chemistry from Charles University in 1961 and a Ph.D. in Biochemistry from the Institute of Organic Chemistry and Biochemistry of the Czechoslovak Academy of Sciences for work on human serum albumin. From 1966-1967 Jarmila was a postdoctoral research associate working on bovine serum albumin with M.J. Hunter at the University of Michigan, Ann Arbor, including the characterization of free sulfhydryl group presence in various forms of albumin. From 1967 to 1973 while in England due to her husband’s job, Jarmila took a break from her lab research career to raise her children Petr and Hana, but still found time to write a major review on the heterogeneity of serum albumin. Jarmila and her family then returned to the United States and she began her long association with the University of Utah in Salt Lake City. Initially, there were a couple of fairly short term postdoctoral research associate positions, first in the lab of W.R. Gray working on tropoelastin (1973-1975), then in the lab of J.D. Andrade working on albumin and biomaterial protein adsorptions (1976-1977), separated by another year in England in between. In the fall of 1977 Jarmila joined the lab group of Dr. James W. Prahl who, together with Dr. Brian F. Tack, had been working for a number of years on purification procedures and other biochemical characterizations of complement proteins C3 and C4 when they were both in the Washington, D.C. area. Their collaboration continued after Dr. Prahl’s arrival in Utah in 1975. A major question of the time in the complement field related to the mechanism of the so-called labile binding site through which a small portion of nascently-activated C3b or C4b could attach the protein to target surfaces, while the rest of the material, which did not immediately find a target, in each case became inactive in the fluid phase. It was towards elucidation of this question that Jarmila joined in on the Prahl lab effort, an effort that ultimately led to the inferential proposal, and then more definitive proof, of the existence of an internal thioester bond in C3 and C4. The key events described below in the discovery of the thioester (or thiol ester, using Jarmila’s preferred nomenclature) are largely gleaned from a first person account that Jarmila wrote in a 1983 New York Academy of Sciences review on the topic.
Just prior to Jarmila Janatova’s arrival in the Prahl lab, Alex Law and Paul Levine published their paradigm-shifting PNAS paper suggesting that a covalent ester bond was formed between nascently-activated C3b and the C3 convertase-bearing target surface, but the mechanism was unknown. The Prahl lab group was aware of some quite old literature showing that nucleophilic reagents such as hydroxylamine and hydrazine, as well as chaotropes and low concentrations of denaturants could render C3 and C4 “hemolytically inactive”, which they understood to mean that the treated molecules could no longer deposit onto convertase-bearing red cells. As part of a systematic effort to determine what biochemical changes in C3 were brought about by these treatments, the group decided to examine whether there was a change in free sulfhydryl content. This is where Jarmila’s vast experience in quantifying the presence of free sulfydryl groups in serum albumin isoforms surely came into play. Jarmila and her colleagues found that compared to native C3, which had no accessible sulfhydryl groups, all of the above treatments resulted in the release of one sulfhydryl group per molecule of C3, as did enzymatic conversion of C3 to C3b. When 14C-iodoacetamide was used to radioalkylate the free SH group, the label was localized to the elastase-generated C3d fragment and it was confirmed to label a cysteine side chain SH. Upon extensive trypsin digestion of hydroxylamine-treated C3, chaotrope-inactivated C3, and enzymatically-produced C3b, all of which were radioalkylated, the label localized in each case to a single tryptic peptide, but whereas in a two dimensional peptide mapping experiment the migration of the peptide from C3b and the chaotrope-treated C3 were identical, that from the hydroxylamine-treated C3 was slightly different, as one would expect if the hydroxylamine nucleophile covalently modified a reactive carbonyl in C3. In a Biochemistry paper in 1980 in which Jarmila Janatova was the first author, the above-described biochemical insights allowed the group to be the first to propose the existence of an internal thiol ester bond in C3 and that this entity mediated the transesterification of nascently-activated C3b to target hydroxyl groups. The very sad part of the story is that just as the last of these ground-breaking experiments were being completed in late June 1979, Dr. James Prahl, died on a mountaineering excursion. Jarmila continued the long distance collaboration with Brian Tack, now in Boston, to solidify their proposal of an internal thioester in C3. Using 14C-methylamine as the nitrogen nucleophile, and through some very clever solid phase thiol chemistry, it was possible to isolate a tryptic peptide containing the free thiol generated by the 14C-methylamine incorporation into C3 and which could then be radioalkylated with 3H-iodoacetic acid. Then through Edman sequencing, the location and residue identities of the labeled amino acids could be determined. This led to the identification of the sequence within the double labeled peptide as Cys*-Gly-Glu-Glx*, where Cys* was 3H-carboxymethycycteine and Glx* was 14C-g-glutamylmethylamide. The close apposition of the liberated sufhydryl and the nucleophile-reactive carbonyl group strongly suggesting that a thioester formed between the thiol of the cysteine and the carbonyl side chain of the glutamyl residue 3 amino acids C-terminal, thus forming a 15-member thiolactone ring entity. The Glx was subsequently identified as Gln when the C3 cDNA sequence became available. In follow-up studies, the Janatova/Tack collaboration went on to provide evidence for the presence of the thioester bond in C4, but not C5.
In 1981 Jarmila was formally appointed as Research Assistant Professor in the Department of Pathology, at the University of Utah, Salt Lake City. She rose to the Associate rank in 1985 and the Adjunct Professor rank in 2004. In 1986 Dr. Janatova became an Academic Member of the Centre for Biopolymers at Interfaces and then she switched her Departmental affiliation to the Department of Bioengineering, University of Utah. With these changes in her appointment, Jarmila’s work also transitioned to having a greater focus on biocompatibility issues of complement with medical biopolymers, as well as studies on the biochemistry of tear proteins and their inhibitors. Together with her graduate students and collaborators, Jarmila had many publications on these topics, as well as others involving methods development of biosensors, but every now and then, Jarmila would still have publications true to her more “traditional” passion of complement protein structure-function relationships. Another noteworthy contribution that Jarmila Janatova made to the complement field was in her role as Secretary on the organizing committee for the very successful ICW XVIII held at the Snowbird Resort, near Salt Lake City, Utah in July 2000. Jarmila Janatova retired from active service at the University of Utah in 2011, retaining the title of Adjunct Professor Emerita in the Department of Bioengineering.
I (DEI) will always remember my first meeting with Jarmila. The date was the morning of July 9, 1979 standing in front of our side-by-side poster boards at the International Congress of Biochemistry meeting in Toronto. Jarmila’s poster on the presence of a single sulfhydryl group in nucleophile-, chaotrope-, or denaturant-treated C3 and C4 had most of the elements of the studies that she and coworkers subsequently published in their landmark papers starting in September 1980. However, the Toronto meeting was a mere week after the tragic death of Dr. James Prahl, and although Jarmila was over the moon with enthusiasm and satisfaction about her scientific results, she was at the same time distressed not only about the loss of her dear friend, colleague and mentor, but also about what the loss of Dr. Prahl, as her scientific advocate, might mean for her career path in Salt Lake City. Because of our overlapping interests in C3 and C4, as well as our biochemistry roots, Jarmila and I bonded very well and we made a habit of sharing at least one meal when we saw each other at conferences in subsequent years. One of these occasions occurred quite early on in our friendship and was at a FASEB meeting in Atlanta, GA. When arranging for a time and place, Jarmila said that dinner would have to be on the early side because she had two tickets to an event that evening and would I like to come. I of course accepted thinking that the tickets might perhaps be for some sporting event that evening. But alas, Jarmila had probably decided that it would be good for me to get some “culturing”, and so she took me to a performance by the Atlanta Symphony Orchestra. I very much enjoyed the performance, but I enjoyed even more watching Jarmila’s enthusiasm about the musical pieces and the quality of their performances. Characteristic of the loud speaking voice with which Jarmila would deliver her complement talks, I can still hear her shouts of BRAVO and BRAVA at the conclusion of the performance. I have very fond memories of our friendship and the many laughs we shared together. I shall very much miss not having the opportunity to share a laugh with her again.
I (AJT) remember Jarmila for many things – her loud laugh making light often of her own disappointment, her love and devotion to her children throughout all stages of their lives, and as an example of how hard it was to be an independent woman in science at that time and particularly without sufficient professional support networks. Many women of that era (who had to make a choice) chose a different path. Jarmila chose to continue to struggle. As further evidence of her love of music and outdoor sports, on a visit to U of U, I took an extra day to visit with her. We went skiing all day and then to the symphony that night. Great day, great evening, great conversations – great friend.
Jarmila was much loved and highly respected by the complement community and she will be greatly missed. Jarmila is survived by her son Petr Janata (neuroscientist), daughter Hana Janatova (musician), brother Josef Tichy, and grandchildren Oliver and Sam Janata. On behalf of all complementologists, we extend our heartfelt condolences to Jarmila’s family.
David E. Isenman Andrea J. Tenner
Department of Biochemistry Depart Molecular Biology & Biochemistry
University of Toronto School of Biological Sciences
Toronto, ON, Canada University of California, Irvine, CA, USA