Another concern related to template based methods is how to meaningfully compare their performance to template-free methods. It is obvious that the query protein structure (for which we want to predict LBS) should be excluded from the template library during evaluation, otherwise the problem is reduced to a simple search. What, then, about very close homologs? To achieve realistic results, authors of eFindSite suggest [45] using sequence identity threshold \( t=40 \% \) (35% in earlier work [52]) and excluding templates with higher sequence identity to the query protein when doing benchmarking predictions. This seems reasonable, albeit any particular choice of threshold t is inevitably arbitrary. For any method other than eFindSite we can find a particular value T for which it will perform roughly the same as eFindSite at \( t=T \).
Our findings must be considered in light of our small sample size. The proportion of bisexual men in our cohort is greater than in comparable cohort analyses from North America [17, 18]. Their profiles and practices may differ from gay men, but the small sample size prevented us from exploring potential subgroup differences based on sexual orientation/identity (gay, bisexual, queer, heterosexual) and sexual behaviours (having sex with men). Participants were recruited from NSPs and open street drug markets, and may not be representative of GBMSM who inject drugs in Melbourne [31] sourcing their drugs and equipment elsewhere. This study was conducted in metropolitan Melbourne, and it is possible that the social ecologies operating in this setting (e.g. drug market characteristics, public policy, network structures, and sociocultural factors) shaped drug practices in unique ways, hindering generalisation. Finally, self-reported variables are invariably subject to social desirability and recall bias. Nonetheless, discrepancies are generally found to be low among people who use drugs, indicating that their self-reports are sufficiently reliable and valid to provide relevant descriptions of drug practices [61, 62].
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